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Gu LH, Fang H, Li FH, Li P, Zhu CX, Zhu JJ, Zhang SJ. Prediction of early hepatic artery thrombosis by intraoperative color Doppler ultrasound in pediatric segmental liver transplantation. Abstract: Early hepatic artery thrombosis (eHAT) after transplantation is associated with a high incidence of graft failure and mortality in pediatric segmental liver transplantation (LT). The evaluation of intraoperative color Doppler ultrasound (CD-US) parameters and their sensitivity and specificity for the prediction of eHAT were important. Pediatric segmental LTs were performed in 49 consecutive patients from October 2006 to December 2010 in our hospital. A total of seven patients (14.3%) experienced eHAT (within one month) after LT. The intraoperative hepatic artery (HA) diameter (p = 0.026), hepatic arterial peak systolic velocity (HAPSV) (p = 0.006), and hepatic artery resistance index (HARI) (p = 0.000) had significant difference between eHAT group and non-eHAT group. Taking a HA diameter

Limited data in animal models suggest that colonic mucosa undergoes adaptive growth following massive small bowel resection (SBR). In vitro data suggest that intestinal cell growth is regulated by reactive oxygen species and redox couples [e.g., glutathione (GSH)/glutathione disulfide (GSSG) and cysteine (Cys)/cystine (CySS) redox]. We investigated the effects of SBR and alterations in redox on colonic growth indexes in rats after either small bowel transection (TX) or 80% midjejunoileal resection (RX). Rats were pair fed ± blockade of endogenous GSH synthesis with buthionine sulfoximine (BSO). Indexes of colonic growth, proliferation, and apoptosis and GSH/GSSG and Cys/CySS redox potentials (Eh) were determined. RX significantly increased colonic crypt depth, number of cells per crypt, and epithelial cell proliferation [crypt cell bromodeoxyuridine (BrdU) incorporation]. Administration of BSO markedly decreased colonic mucosal GSH, GSSG, and Cys concentrations in both TX and RX groups, with a resultant oxidation of GSH/GSSG and Cys/CySS Eh. BSO did not alter colonic crypt cell apoptosis but significantly increased all colonic mucosal growth indexes (crypt depth, cells/crypt, and BrdU incorporation) in both TX and RX groups in a time- and dose-dependent manner. BSO significantly decreased plasma GSH and GSSG, oxidized GSH/GSSG Eh, and increased plasma Cys and CySS concentrations. Collectively, these data provide in vivo evidence indicating that oxidized colonic mucosal redox status stimulates colonic mucosal growth in rats. The data also suggest that GSH is required to maintain normal colonic and plasma Cys/CySS homeostasis in these animal models.

Obesity-related glucose intolerance is a function of hepatic (homeostatic model assessment-insulin resistance [HOMA-IR]) and peripheral insulin resistance (Si) and β-cell dysfunction. We determined relationships between changes in these measures, visceral (VAT) and subcutaneous (SAT) adipose tissue, and systemic adipocytokine biomarkers 1 and 6 months after surgical weight loss. HOMA-IR decreased significantly (−50%) from baseline by 1 month and decreased further (−67%) by 6 months, and Si was improved by 6 months (2.3-fold) weight loss. Plasma concentrations of leptin decreased and adiponectin increased significantly by 1 month, and decreases in interleukin-6, C-reactive protein (CRP), and tumor necrosis factor-α were observed at 6 months of weight loss. Longitudinal decreases in CRP (r = −0.53, P < 0.05) were associated with increases in Si, and decreases in HOMA-IR were related to increases in adiponectin (r = −0.37, P < 0.05). Decreases in VAT were more strongly related to increases in adiponectin and decreases in CRP than were changes in general adiposity or SAT. Thus, in severely obese women, specific loss of VAT leads to acute improvements in hepatic insulin sensitivity mediated by increases in adiponectin and in peripheral insulin sensitivity mediated by decreases in CRP.

Glutathione (GSH) concentration affects cell proliferation and apoptosis in intestinal and other cell lines in vitro. However, in vivo data on gut mucosal GSH redox status and cell turnover are limited. We investigated the effect of altered GSH redox status on the ileal mucosa in a rat model of short bowel syndrome following massive small bowel resection (SBR). Rats underwent 80% mid-jejunoileal resection (RX) or small bowel transection (TX; as operative controls), with administration of either saline or o, L-buthionine-sulfoximine (BSO), a specific inhibitor of cellular GSH synthesis. Ileal mucosal redox, morphology, and indices of cell proliferation and apoptosis were determined at different days after surgery. Ileal GSH redox status was assessed by GSH and GSH disulfide (GSSG) concentrations and the redox potential of GSH/GSSG (E h ). Ileal lipid peroxidation [free malondialdehyde (MDA)] was measured as an index of lipid peroxidation. BSO markedly decreased ileal mucosal GSH, oxidized GSH/GSSG E h , and increased MDA content without inducing morphological damage as assessed by light or electron microscopy. As expected, SBR stimulated adaptive growth of ileal villus height and total mucosal height at 7 d after surgery, but this response was unaffected by BSO treatment despite a modest increase in crypt cell apoptosis. Ileal cell proliferation (crypt cell bromodeoxyuridine incorporation) increased at 2 d after SBR but was unaffected by BSO. Collectively, our in vivo data show that marked depletion of ileal GSH and oxidation of the GSH redox pool does not alter indices of ileal epithelial proliferation or SBR-induced ileal mucosal adaptive growth.

Sensitive biomarkers for intestinal absorptive function would be clinically useful in short bowel syndrome (SBS). Citrulline (Cit) is a product of the metabolism of glutamine (Gln) and derived amino acids by enterocytes. Cit is produced almost exclusively by the gut, which is also a major site of Gln metabolism. The goals of this study were to examine whether plasma Cit and Gln concentrations are biomarkers of residual small intestinal length and nutrient absorptive functions in adult SBS patients followed prospectively. We studied 24 stable adults with severe SBS receiving chronic parenteral nutrition (PN) in a double-blind, randomized trial of individualized dietary modification +/- recombinant human growth hormone (GH). During a baseline week, intestinal absorption studies (% absorption of fluid, kcal, nitrogen, fat, carbohydrate, sodium, phosphorus, and magnesium) were performed and concomitant plasma Cit and Gln concentrations determined. Individualized dietary modification and treatment with subcutaneous injection of placebo (n = 9) or GH (0.1 mg/kg daily x 21 days, then 3 times/week; n = 15) were then begun. PN weaning was initiated after week 4 and continued as tolerated for 24 weeks. Repeat plasma amino acid determination and nutrient absorption studies were performed at weeks 4 and 12. Residual small bowel length at baseline was positively correlated with baseline plasma Cit (r = 0.467; p = .028). However, no significant correlations between absolute Cit or Gln concentrations and the percent absorption of nutrient substrates at any time point were observed. Similarly, no correlation between the change in Cit or GLN concentration and the change in % nutrient absorption was observed (baseline vs weeks 4 and 12, respectively). By weeks 12 and 24, 7 and 13 subjects were weaned completely from PN, respectively. However, baseline plasma Cit or Gln did not predict PN weaning at these time points. We concluded that plasma Cit (but not Gln) concentrations appeared to be an indicator of small intestinal length in adult SBS. However, neither plasma Cit nor Gln was a biomarker for intestinal absorptive function in this cohort of patients with SBS.

Patients with short bowel syndrome (SBS) have a high prevalence of metabolic bone disease due to nutrient malabsorption and potential effects of parenteral nutrition (PN). Human growth hormone (hGH) has been shown in some studies to have anabolic effects on bone, but hGH effects on bone in patients with SBS are unknown.Adults with PN-dependent SBS underwent a 7-day period of baseline studies while receiving usual oral diet and PN and then began receiving modified diets designed to improve nutrient absorption and daily oral calcium/vitamin D supplements (1500 mg elemental calcium and 600 IU vitamin D, respectively). Subjects were randomized to receive in a double-blind manner either subcutaneous (sc) saline placebo as the control or hGH (0.1 mg/kg/d for 3 weeks, then 0.1 mg/kg 3 days a week for 8 subsequent weeks). Open-label hGH was given from week 13 to week 24 in subjects who required PN after completion of the 12-week double-blind phase. Markers of bone turnover (serum osteocalcin and urinary N-telopeptide [NTX]), vitamin D nutriture (serum calcium, 25-hydroxyvitamin D [25-OH D] and parathyroid hormone [PTH] concentrations), and intestinal calcium absorption were measured at baseline and at weeks 4 and 12. Dual x-ray absorptiometry (DXA) of the hip and spine was performed to determine bone mineral density (BMD) at baseline and weeks 12 and 24.The majority of subjects in each group exhibited evidence of vitamin D deficiency at baseline (25-OH D levels30 ng/mL; 78% and 79% of control and hGH-treated subjects, respectively). Subjects treated with hGH demonstrated a significant increase from baseline in serum osteocalcin levels at 12 weeks (+62%; p.05). The levels of NTX were increased over time in the hGH-treated group; however, this did not reach statistical significance. Both NTX and osteocalcin remained unchanged in control subjects. BMD of the spine and total hip was unchanged in subjects treated with placebo or hGH at 24 weeks. However, femoral neck BMD was slightly but significantly decreased in the placebo group at this time point but remained unchanged from baseline in the hGH-treated subjects.hGH therapy significantly increased markers of bone turnover during the initial 3 months of therapy and stabilized femoral neck bone mass over a 6-month period in patients with severe SBS undergoing intestinal rehabilitation.

Low molecular weight thiol/disulfide redox pools are dependent upon extracellular cysteine (Cys) availability. We determined whether dietary sulfur amino acid (SAA) deficiency induces oxidative stress in vivo, as determined by redox state of major thiol/disulfide couples in plasma [Cys/cystine (CySS)] and intestinal mucosa [glutathione (GSH)/glutathione disulfide (GSSG)]. Rats were fed isocaloric, isonitrogenous semipurified diets: either SAA-adequate (control), SAA-deficient, or SAA-supplemented, pair-fed to intake of the SAA-deficient group. Reference rats consumed standard rat food ad libitum. After 7 d, plasma and gut mucosal samples were analyzed for Cys, CySS, GSH and GSSG, and the redox potentials of Cys/CySS and GSH/GSSG were determined. Mean daily food intake in the pair-fed rats was similar (approximately one-half of reference-rat intake). Body weight decreased in all pair-fed groups, but rats fed the SAA-deficient diet lost significantly more body weight. Dietary SAA deficiency decreased GSH concentrations in both plasma and gut mucosa, increased plasma GSSG, and oxidized plasma and gut mucosal GSH/GSSG redox and plasma Cys/CySS redox. SAA supplementation resulted in a more reducing plasma Cys/CySS redox potential. Reference rats exhibited similar tissue and plasma GSH/GSSG redox as rats that ate semipurified SAA-adequate rat food, which provided similar net SAA intake. Our in vivo data show that inadequate dietary SAA intake oxidizes the thiol/disulfide redox status in rat-gut mucosa and plasma. Such oxidation of redox pools is associated with oxidative stress and the onset or progression of several pathological conditions. Thus, dietary SAA deficiency could contribute to the progression of disease by causing an oxidation of these components.

Background Massive small-bowel resection (SBR) increases adaptive growth of residual intestine in animal models of short-bowel syndrome (SBS). Pyrimidine nucleotides are critical for DNA and RNA synthesis, but no previous study has evaluated whether supplementation of pyrimidines or their precursors in the diet enhances adaptive gut growth after SBR. This study determined growth responses in jejunal mucosa after 7 days of dietary supplementation with uracil, or its precursor, orotate, after massive SBR in rats. Methods Sprague-Dawley rats ( approximately 200 g) underwent 80% jejunoileal resection (RX) or ileal transection (TX; control). Rats were pair-fed a purified (AIN-93G) powdered diet supplemented with or without 1% (wt/wt) orotate or uracil until killing at 7 days postsurgery. Defined jejunal segments were obtained for analysis of mucosal villus height (VH), crypt depth (CD), total mucosal height, bromodeoxyuridine (BrdU) incorporation, an index of cell proliferation, and full-thickness DNA and protein content as measures of intestinal adaptive growth. Results Jejunal VH increased significantly with SBR, as expected, and orotate further stimulated this response. Jejunal CD and total mucosal height increased significantly with both orotate and uracil supplementation compared with resected animals receiving standard diet. Orotate administration also increased jejunal DNA content compared with the increase observed with SBR alone. Finally, orotate, but not uracil, supplementation increased BrdU incorporation compared with resected rats fed standard or uracil-supplemented diet after SBR. Conclusions Supplementation of oral diet with the pyrimidine precursor orotate and uracil stimulated adaptive jejunal growth after massive SBR in rats. Dietary orotate had more potent growth-stimulatory effects than uracil in this animal model. Dietary supplementation with orotate and uracil represents a novel nutrition approach to enhance small-bowel mucosal adaptive growth and absorptive capacity in SBS.

Wnts are morphogens with well recognized functions during embryogenesis. Aberrant Wnt signaling has been demonstrated to be important in colorectal carcinogenesis. However, the role of Wnt in regulating normal intestinal epithelial cell proliferation is not well established. Here we determine that Wnt11 is expressed throughout the mouse intestinal tract including the epithelial cells. Conditioned media from Wnt11-secreting cells stimulated proliferation and migration of IEC6 intestinal epithelial cells. Co-culture of Wnt11-secreting cells with IEC6 cells resulted in morphological transformation of the latter as evidenced by the formation of foci, a condition also accomplished by stable transfection of IEC6 with a Wnt11-expressing construct. Treatment of IEC6 cells with Wnt11 conditioned media failed to induce nuclear translocation of beta-catenin but led to increased activities of protein kinase C and Ca(2+)/calmodulin-dependent protein kinase II. Inhibition of protein kinase C resulted in a decreased ability of Wnt11 to induce foci formation in IEC6 cells. Finally, E-cadherin was redistributed in Wnt11-treated IEC6 cells, resulting in diminished E-cadherin-mediated cell-cell contact. We conclude that Wnt11 stimulates proliferation, migration, cytoskeletal rearrangement, and contact-independent growth of IEC6 cells by a beta-catenin-independent mechanism. These findings may help understand the molecular mechanisms that regulate proliferation and migration of intestinal epithelial cells.

Azathioprine is commonly prescribed for autoimmune hepatitis and inflammatory bowel disease. An acute gastroenteritis-like syndrome has been ascribed to azathioprine use, but chronic diarrhea has not. We report a patient with autoimmune hepatitis who developed severe small-bowel villus atrophy and chronic diarrhea after azathioprine was initiated (50 mg/day). We present a case report of a patient followed up prospectively. Duodenal mucosal histology and expression of brush border enzyme dipeptidyl peptidase IV and peptide transporter PepT1 messenger RNA levels were determined before and after azathioprine discontinuation. Chronic diarrhea developed several weeks after the initiation of azathioprine and resulted in micronutrient depletion and severe protein-calorie malnutrition, which was unresponsive to oral pancreatic enzyme therapy or a gluten-free diet. Severe malabsorption required parenteral nutrition support for longer than 1.5 years; this was complicated by unstable blood glucose control, acute calculous cholecystitis, catheter sepsis, and severe venous thrombosis. When the temporal association between azathioprine and diarrhea was identified, the drug was tapered while the patient consumed an unrestricted diet. Within 2 weeks after azathioprine was discontinued, diarrhea had completely resolved, and parenteral nutrition was discontinued. Mucosal biopsies obtained before and 4 months after azathioprine discontinuation showed complete reversal of severe duodenal villus atrophy and marked up-regulation of mucosal dipeptidyl peptidase IV and PepT1 messenger RNA. The patient has subsequently maintained normal liver function tests on low-dose prednisone alone, with normal stools and stable nutritional status for longer than 4 years. Azathioprine can induce severe small-bowel villus atrophy, diarrhea, and malabsorption that is reversible with drug discontinuation.

The trefoil factor family peptides TFF1, TFF2, and TFF3 are important for gut mucosal protection and restitution. Keratinocyte growth factor (KGF) stimulates proliferation and differentiation of epithelial cells with potent effects on goblet cells. To investigate interactions between food intake and KGF, rats were fed ad libitum (control), fasted for 72 h, or fasted for 72 h and then refed for 72 h with or without KGF (3 mg · kg−1· day−1). With fasting, goblet cell number in duodenum increased, TFF3 mRNA in duodenum and jejunum decreased, and TFF3 protein did not change or increased. KGF during fasting stimulated colonic growth, normalized TFF3 mRNA in duodenum and jejunum, and broadly upregulated gut goblet cell number and TFF3 protein expression. With fasting-refeeding, KGF increased small bowel and colonic mucosal growth, goblet cell number, and TFF3 protein but had variable effects on TFF3 mRNA. KGF induced TFF2 mRNA and protein in duodenum and jejunum with both nutritional regimens. We conclude that nutrient availability modifies rat intestinal goblet cell number, TFF3 mRNA, and the gut-trophic effects of KGF in a region-specific manner. KGF enhances TFF2 expression in proximal small bowel and increases goblet cell number and TFF3 protein content throughout the intestine independent of food intake.

Redox mechanisms function in regulation of cell growth, and variation in redox state of plasma thiol/disulfide couples occurs in various physiologic conditions, including diabetes, chemotherapy, and aging. The present study was designed to determine whether a systematic variation in extracellular thiol/disulfide redox state (E(h)) over a range (0 mV to -150 mV) that occurs in human plasma altered proliferation of cultured cells. Experiments were performed with a human colon carcinoma cell line (Caco2), which grows slowly in the absence of serum and responds to peptide growth factors with increased rate of cell division. The extracellular redox states were established by varying concentrations of cysteine and cystine, maintaining constant pool size in terms of cysteine equivalents. Incorporation of 5-bromo-2-deoxyuridine (BrdU) was used to measure DNA synthesis and was lowest at the most oxidized extracellular E(h) (0 mV). Incorporation increased as a function of redox state, attaining a 100% higher value at the most reduced condition (-150 mV). Addition of insulin-like growth factor-1 (IGF-1) or epidermal growth factor (EGF) increased the rate of BrdU incorporation at more oxidizing redox conditions (0 to -80 mV) but had no effect at -150 mV. Cellular GSH was not significantly affected by variation in extracellular E(h). In the absence of growth factors, extracellular E(h) values were largely maintained for 24 h. However, IGF-1 or EGF stimulated a change in extracellular redox to values similar to that for cysteine/cystine redox in plasma of young, healthy individuals. The results show that extracellular thiol/disulfide redox state modulates cell proliferation rate and that this control interacts with growth factor signaling apparently independently of cellular glutathione.

There is an acute need for effective therapy for inflammatory bowel disease (IBD), particularly at the level of repair of the damaged epithelium. We evaluated the efficacy of recombinant human keratinocyte growth factor (rHuKGF) in both the dextran sodium sulfate (DSS) and the CD4+CD45RBHiT cell transfer models of IBD. Disease was induced either by the ad libitum administration to normal mice of 4% DSS in the drinking water or by the injection of 4 × 105CD4+CD45RBHiT cells into immunodeficient scid/scid mice. rHuKGF was administered by subcutaneous injection at doses of 1.0 or 3.0 mg/kg in both preventative and therapeutic regimens during both studies. rHuKGF significantly improved survival and body weight loss in the DSS model in both preventative and therapeutic dosing regimens. It also improved diarrhea, hematochezia, and hematological parameters, as well as large intestine histopathology. In the T cell transfer model, rHuKGF improved body weight loss, diarrhea, and levels of serum amyloid A, as well as large intestine histopathology. In both models of IBD, the colonic levels of intestinal trefoil factor (ITF) were elevated by the disease state and further elevated by treatment with rHuKGF. These data suggest that rHuKGF may prove useful in the clinical management of IBD and its effects are likely mediated by its ability to locally increase the levels of ITF.

Background. Reactive oxygen species (ROS) and glutathione (GSH) depletion contribute to organ injury after bone marrow transplantation (BMT). Keratinocyte growth factor (KGF) ameliorates graft-versus-host disease (GVHD)-associated organ injury in murine BMT models. Methods. B10.BR mice received total body irradiation (TBI; day −1) ± cyclophosphamide (Cy; 120 mg/kg/day i.p., days −3 and −2), then were transplanted on day 0 with C57BL/6 bone marrow + spleen cells as a source of GVHD-causing T cells. KGF (5 mg/kg/day subcutaneously [s.c.]) or saline was given on days −6, −5, and −4. Lung and liver GSH and oxidized GSH disulfide (GSSG) levels were measured on days 0 and 5 and glutathione redox potential (Eh) calculated. Organ malondialdehyde (MDA) was determined on day 5 as an index of ROS-mediated lipid peroxidation. Results. In lung, TBI+BMT oxidized GSH Eh and increased MDA. Cy further oxidized lung GSH Eh. In liver, neither BMT regimen altered GSH redox status or MDA. KGF prevented the decrease in lung GSH after TBI+Cy and decreased lung MDA after both TBI and TBI+Cy. KGF increased liver GSH levels and GSH Eh after TBI and GSH Eh after TBI+Cy. Conclusions. In murine allogeneic BMT, TBI oxidizes the lung GSH redox pool and Cy exacerbates this response by 5 days post-BMT. In contrast, liver GSH redox status is maintained under these experimental conditions. KGF treatment attenuates the Cy-induced decrease in lung GSH, decreases post-BMT lung lipid peroxidation, and improves liver GSH redox indices. KGF may have a therapeutic role to prevent or attenuate GSH depletion and ROS-mediated organ injury in BMT.

Malnutrition is associated with increased reactive oxygen species (ROS) formation and depletion of the critical antioxidant glutathione (GSH) in the intestine. The malnutrition-induced decrease in gut GSH levels is prevented by recombinant keratinocyte growth factor (KGF) administration. We investigated whether enzymes that are induced by oxidants and modulate tissue GSH supply are regulated by enteral nutrients or KGF at the messenger RNA (mRNA) level.Adult rats were fasted for 3 days alone or fasted for 3 days then refed ad libitum. In a second model, rats were fasted for 3 days and then refed ad libitum or 25% of ad libitum intake with daily intraperitoneal saline or recombinant KGF (5 mg/kg/d) for 3 subsequent days. mRNA levels for gamma-glutamylcysteine synthetase (gamma-GCS), gamma-glutamyl transpeptidase (gamma-GT), glutathione-S-transferase Ya-subunit, gastrointestinal glutathione peroxidase (GI-GPx), and non-selenium-dependent glutathione peroxidase (ns-GPx) were determined in ileum and colon by ribonuclease protection assay.Fasting increased ileal gamma-GCS, ns-GPx, and glutathione-S-transferase mRNAs (by 36%, 165%, and 130% of controls) and decreased GI-GPx mRNA (to 55% of controls). In the colon, mRNAs for GSH-related enzymes were unchanged by fasting or refeeding. Prolonged enteral nutrient restriction (25% refeeding after a 3-day fast) increased gamma-GCS and glutathione-S-transferase mRNAs (by270% of controls), decreased GI-GPx mRNA (to50% of controls) in ileum and colon and increased ns-GPx mRNA (by 180%) in colon. KGF treatment increased ns-GPx mRNA in the ileum and colon and glutathione-S-transferase mRNA in the colon (by200% of controls).Enteral nutrient intake regulates GSH-related enzyme mRNA levels in the intestine, which may contribute to the decrease in mucosal GSH during malnutrition. Increased ns-GPx and glutathione-S-transferase mRNA levels during malnutrition and with KGF administration may increase detoxifying functions in the gut under these conditions.

Several lines of evidence demonstrate that general nutritional status, specific nutrients (eg, zinc, glutamine), and certain trophic growth factors (eg, growth hormone, insulin-like growth factor I, keratinocyte growth factor, and glucagon-like peptide-2) have important interactions relevant for intestinal growth and function. Adequate nutritional status is critical for endogenous growth factor synthesis in the gut and other tissues and is an important mediator of organ responsiveness to exogenous growth factor administration. Both endogenously synthesized and exogenously administered growth factors upregulate nutrient uptake and utilization by gut mucosa, skeletal muscle, and other organs. Emerging data from both animal and human studies indicate that combinations of selected growth factors and specific nutrients may improve the growth, adaptation, and repair of the intestinal mucosa. Additional studies to determine basic mechanisms of nutrient-growth factor interactions and the safety and efficacy of treatment with combinations of specific nutrients and recombinant growth factors are needed. Results of these investigations should define new methods for support of the intestinal tract during short bowel syndrome (SBS), catabolic illness, and malnutrition.

BACKGROUND Keratinocyte growth factor (KGF) induces proliferation of gut epithelium in rat models, but KGF-nutrient interactions have not been studied. An experimental model of fasting-induced gut atrophy followed by different levels of enteral refeeding was used to investigate the influence of nutrient availability on the gut-trophic effects of exogenous KGF. METHODS After a 3-day fast, rats were enterally refed either ad libitum or at 25% of ad libitum intake for 3 subsequent days. Either intraperitoneal KGF (5 mg/kg/d) or saline was given in each dietary regimen. Wet weight, DNA, and protein content were measured as indices of full-thickness cellularity in duodenum, jejunum, ileum, and colon. Villus height in small bowel segments and crypt depth in all gut tissues were measured as specific indices of mucosal growth. RESULTS Refeeding at 25% of ad libitum intake significantly decreased full-thickness cellularity and mucosal growth indices in duodenum, jejunum, and ileum. In the colon, only protein content fell significantly and crypt depth was maintained. KGF administration during 25% refeeding did not alter full-thickness indices in any small bowel segment or affect jejunal mucosal growth. In contrast, KGF normalized duodenal villus height (p < .01) and duodenal and ileal crypt depth (p < .05) only in the 25%-refed model. KGF significantly increased ileal villus height in both ad libitum and 25%-refed rats (by 43% and 48%, respectively, p < .05) and markedly increased colonic cellularity and mucosal crypt depth with both levels of refeeding (p < .01). CONCLUSIONS Rat small bowel growth is more sensitive than colon to the level of enteral refeeding after a 3-day fast. KGF administration does not affect jejunal growth, but specifically prevents atrophy of duodenal and ileal mucosa during hypocaloric, hyponitrogenous refeeding. In ileum and colon, some KGF-mediated growth responses are independent of the level of enteral refeeding. Thus gut-trophic effects of KGF and KGF interactions with the level of nutrient intake are tissue-specific.

Early hepatic artery thrombosis (eHAT) after transplantation is associated with a high incidence of graft failure and mortality in pediatric segmental liver transplantation (LT). The evaluation of intraoperative color Doppler ultrasound (CD-US) parameters and their sensitivity and specificity for the prediction of eHAT were important. Pediatric segmental LTs were performed in 49 consecutive patients from October 2006 to December 2010 in our hospital. A total of seven patients (14.3%) experienced eHAT (within one month) after LT. The intraoperative hepatic artery (HA) diameter (p = 0.026), hepatic arterial peak systolic velocity (HAPSV) (p = 0.006), and hepatic artery resistance index (HARI) (p = 0.000) had significant difference between eHAT group and non-eHAT group. Taking a HA diameter2 mm, a HAPSV of40 cm/s and a HARI of0.6 as threshold to predict eHAT, the sensitivity and specificity were 85.7%, 85.7%, 85.7%, and 61.9%, 76.2%, 88.1%, respectively. A HARI of0.6 was shown to be the most sensitive and specific single parameter for predicting eHAT.

Glutathione (GSH) concentration affects cell proliferation and apoptosis in intestinal and other cell lines in vitro. However, in vivo data on gut mucosal GSH redox status and cell turnover are limited. We investigated the effect of altered GSH redox status on the ileal mucosa in a rat model of short bowel syndrome following massive small bowel resection (SBR). Rats underwent 80% mid-jejunoileal resection (RX) or small bowel transection (TX; as operative controls), with administration of either saline or D, L-buthionine-sulfoximine (BSO), a specific inhibitor of cellular GSH synthesis. Ileal mucosal redox, morphology, and indices of cell proliferation and apoptosis were determined at different days after surgery. Ileal GSH redox status was assessed by GSH and GSH disulfide (GSSG) concentrations and the redox potential of GSH/GSSG (Eh). Ileal lipid peroxidation [free malondialdehyde (MDA)] was measured as an index of lipid peroxidation. BSO markedly decreased ileal mucosal GSH, oxidized GSH/GSSG Eh, and increased MDA content without inducing morphological damage as assessed by light or electron microscopy. As expected, SBR stimulated adaptive growth of ileal villus height and total mucosal height at 7 d after surgery, but this response was unaffected by BSO treatment despite a modest increase in crypt cell apoptosis. Ileal cell proliferation (crypt cell bromodeoxyuridine incorporation) increased at 2 d after SBR but was unaffected by BSO. Collectively, our in vivo data show that marked depletion of ileal GSH and oxidation of the GSH redox pool does not alter indices of ileal epithelial proliferation or SBR-induced ileal mucosal adaptive growth.

Background Administration of specific growth factors exert gut-trophic effects in animal models of massive small bowel resection (SBR); however, little comparative data are available. Our aim was to compare effects of a human glucagon-like peptide-2 (GLP-2) analog, recombinant growth hormone (GH) and recombinant keratinocyte growth factor (KGF) on jejunal, ileal, and colonic growth and functional indices after 80% SBR in rats. Methods Thirty-seven male rats underwent small bowel transection (sham operation) with s.c. saline administration (control; Tx-S; n = 7) or 80% midjejuno-ileal resection (Rx) and treatment with either s.c. saline (Rx-S, n = 7), GLP-2 at 0.2 mg/kg/d (Rx-GLP-2; n = 8), GH at 3.0 mg/kg/d (Rx-GH; n = 8), or KGF at 3.0 mg/kg/d (Rx-KGF; n = 7) for 7 days. All groups were pair-fed to intake of Rx-S rats. Gut mucosal cell growth indices (wet weight, DNA and protein content, villus height, crypt depth, and total mucosal height) were measured. Expression of the cytoprotective trefoil peptide TFF3 was determined by Western blot. Gut mucosal concentrations of the tripeptide glutathione (L-glutamyl-L-cysteinyl-glycine) and glutathione disulfide (GSSG) were measured by high-performance liquid chromatography and the glutathione/GSSG ratio calculated. Results SBR increased adaptive growth indices in jejunal, ileal, and colonic mucosa. GLP-2 treatment increased jejunal villus height and jejunal total mucosal height compared with effects of resection alone or resection with GH or KGF treatment. Both GH and KGF modestly increased colonic crypt depth after SBR. SBR did not affect small bowel or colonic goblet cell number or TFF3 expression; however, goblet cell number and TFF3 expression in both small bowel and colon were markedly up-regulated by KGF treatment and unaffected by GLP-2 and GH. SBR oxidized the ileal and colonic mucosal glutathione/GSSG redox pools. GLP-2 treatment after SBR increased the glutathione/GSSG ratio in jejunum, whereas KGF had an intermediate effect. In addition, GLP-2 (but not GH or KGF) prevented the SBR-induced oxidation of the glutathione/GSSG pools in both ileum and colon. Conclusions GLP-2 exerts superior trophic effects on jejunal growth and also improves mucosal glutathione redox status throughout the bowel after massive SBR in rats. Both GH and KGF increase colonic mucosal growth in this model. KGF alone potently increases gut mucosal goblet cell number and expression of the cytoprotective trefoil peptide TFF3. The differential effects of GLP-2, GH and KGF administration in this model of short bowel syndrome suggest that individual therapy with these growth factors may not be an adequate strategy to maximally improve adaptive gut mucosal growth and cytoprotection after massive small intestinal resection. Future research should address the use of these agents in combination in short bowel syndrome.

Glutamine (Gln) and keratinocyte growth factor (KGF) each stimulate intestinal epithelial cell growth, but regulatory mechanisms are not well understood. We determined whether Gln and KGF alter intra- and extracellular thiol/disulfide redox pools in Caco-2 cells cultured in oxidizing or reducing cell medium and whether such redox variations are a determinant of proliferative responses to these agents. Cells were cultured over a physiological range of oxidizing to reducing extracellular thiol/disulfide redox (Eh) conditions, obtained by varying cysteine (Cys) and cystine (CySS) concentrations in cell medium. Cell proliferation was determined by 5-bromo-2-deoxyuridine (BrdU) incorporation. Gln (10 mmol/l) or KGF (10 μg/l) did not alter BrdU incorporation at reducing Eh (-131 to -150 mV), but significantly increased incorporation at more oxidizing Eh (Gln at 0 to -109 mV; KGF at -46 to -80 mV). Cellular glutathione/glutathione disulfide (GSH/GSSG) Eh was unaffected by Gln, KGF, or variations in extracellular Cys/CySS Eh. Control cells largely maintained extracellular Eh at initial values after 24 h (-36 to -136 mV). However, extracellular Eh shifted toward a narrow physiological range with Gln and KGF treatment (Gln -56 to -88 mV and KGF -76 to -92 mV, respectively; P < 0.05 vs. control). The results indicate that thiol/disulfide redox state in the extracellular milieu is an important determinant of Caco-2 cell proliferation induced by Gln and KGF, that this control is independent of intracellular GSH redox status, and that both Gln and KGF enhance the capability of Caco-2 cells to modulate extremes of extracellular redox.

Cellular redox, maintained by the glutathione (GSH)- and thioredoxin (Trx)-dependent systems, has been implicated in the regulation of a variety of biological processes. The redox state of the GSH system becomes oxidized when cells are induced to differentiate by chemical agents. The aim of this study was to determine the redox state of cellular GSH/glutathione disulfide (GSH/GSSG) and Trx as a consequence of progression from proliferation to contact inhibition and spontaneous differentiation in colon carcinoma (Caco-2) cells. Results showed a significant decrease in GSH concentration, accompanied by a 40-mV oxidation of the cellular GSH/GSSG redox state and a 28-mV oxidation of the extracellular cysteine/cystine redox state in association with confluency and increase in differentiation markers. The redox state of Trx did not change. Thus the two central cellular antioxidant and redox-regulating systems (GSH and Trx) were independently controlled. According to the Nernst equation, a 30-mV oxidation is associated with a 10-fold change in the reduced/oxidized ratio of a redox-sensitive dithiol motif. Therefore, the measured 40-mV oxidation of the cellular GSH/GSSG couple or the 28-mV oxidation of the extracellular cysteine/cystine couple should be sufficient to function in signaling or regulation of differentiation in Caco-2 cells.

Background: Intestinal adaptation after massive bowel resection in animal models is characterized by increased gut-mucosal growth and expression of nutrient transporters. Few data about these indexes exist in humans with short-bowel syndrome (SBS). Objective: The objective was to compare small-bowel and colonic mucosal growth and expression of the peptide transporter PepT1 in adults with or without SBS. Design: Mucosal biopsy specimens were obtained from the small bowel and colon of 33 control subjects with intact intestine and from 13 SBS patients dependent on parenteral nutrition because of chronic malabsorption. Gut-mucosal crypt depth, villus height, and villus width were measured, and expression of PepT1 was determined by Northern blotting, in situ hybridization, and immunohistochemistry. Results: The indexes of small-bowel and colonic mucosal growth were not significantly different between the 2 groups. PepT1 expression was high in the apical region of duodenal, jejunal, and ileal villus epithelial cells; low in absorptive colonocytes; and not significantly different in the distal small intestine of the 2 groups. However, the abundance of PepTI mRNA in the colon of SBS patients was more than 5-fold that in control subjects (P < 0.01). Conclusions: Gut adaptation in SBS patients does not appear to involve an increase in gut-mucosal crypt depth or villus size. PepT1 is abundant along the small-bowel brush border in humans; expression in the colon indicates that the large intestine has a mechanism for luminal di- and tripeptide transport. Up-regulation of colonic PepT1 in SBS may adaptively improve accrual of malabsorbed di- and tripeptides, independent of changes in the mucosal surface area.

Sucessful intestinal adaptation after massive enterectomy is dependent on increased efficiency of nutrient transport. However, midgut resection (MGR) in rabbits induces an initial decrease in sodium-dependent brush border neutral amino acid transport, whereas parenteral epidermal growth factor (EGF) and growth hormone (GH) reverse this downregulation. We investigated intestinal amino acid transporter B0 (ATB0) and oligopeptide transporter 1 (PEPT 1) mRNA expression after resection and in response to EGF and/or GH.Rabbits underwent anesthesia alone (control) or proximal, midgut, and distal resections. Full-thickness intestine was harvested from all groups on postoperative day (POD) 7, and on POD 14 from control and MGR rabbits. A second group of MGR rabbits received EGF and/or GH for 7 days, beginning 7 days after resection. ATB0 and PEPT 1 mRNA levels were determined by Northern blot analysis.In control animals, ileal ATB0 mRNA abundance was three times higher than jejunal mRNA, whereas PEPT 1 mRNA expression was similar. By 7 and 14 days after MGR, jejunal ATB0 mRNA abundance was decreased by 50% vs control jejunum. A 50% decrease in jejunal PEPT 1 message was delayed until 14 days after MGR. Treatment with EGF plus GH did not alter ATB0 mRNA expression but doubled PEPT 1 mRNA in the jejunum.The site of resection, time postresection, and growth factors treatment differentially influence ATB0 and PEPT 1 mRNA expression. Enhanced sodium-dependent brush border neutral amino acid transport with GH plus EGF administration is independent of increased ATB0 mRNA expression in rabbit small intestine after enterectomy.

Malnutrition decreases tissue levels of glutathione (GSH), a major endogenous antioxidant that detoxifies reactive oxygen species and promotes cell growth. This study determined the effects of the gut trophic peptide keratinocyte growth factor (KGF) on intestinal mucosal GSH concentrations and redox state in malnourished rats. Adult rats were food-deprived for 3 d, then consumed food ad libitum or 25% of ad libitum intake for 3 d with daily intraperitoneal administration of saline or KGF (5 mg.kg -1 .d -1 ). Mucosal GSH and glutathione disulfide (GSSG) concentrations, crypt depth and total mucosal height were measured in the jejunum, ileum and colon. In the 25% of ad libitum-refed, saline-treated group, mucosal GSH was lower in all gut tissues (42% in jejunum, 38% in ileum, and 57% in colon), and the GSH/GSSG ratio was lower in the jejunum and ileum compared to that in the ad libitum-refed controls. KGF treatment with ad libitum refeeding increased GSH/GSSG in the jejunum, ileum and colon. Furthermore, in 25% of ad libitum refeeding, KGF normalized jejunal, ileal and colonic mucosal GSH content and significantly increased the mucosal GSH/GSSG ratio relative to rats treated with saline. Increased crypt depth and total mucosal height induced by KGF and feeding could be explained in part by increased mucosal GSH content. KGF treatment improved gut mucosal glutathione redox state in malnourished, refed rats. These data provide evidence that gut trophic hormones and food intake may independently support gut mucosal glutathione antioxidant capacity during nutritional repletion.

Background. After massive enterectomy (ME), remnant intestine undergoes compensatory adaptation. Epidermal growth factor (EGF) and human growth hormone (hGH) have each been shown to enhance total length small intestine nutrient transport after ME. This study aims to determine the differential effects of EGF and hGH on proximal and distal small intestinal remnants after ME. Methods. New Zealand white rabbits underwent 70% mid-jejunoileal resection. After 1 week, animals received hGH (0.2 mg/kg/day), EGF (1.5 μg/kg/hr), hGH + EGF, or vehicle (equal volume) for 7 days. Sodium-dependent uptake of glucose, glutamine, alanine, leucine, and arginine into brush border membrane vesicles was quantitated. Serum insulin-like growth factor-I concentrations as well as proximal and distal villus and microvillus heights were measured. IGF binding protein-3 and -4 mRNA expression was determined in full-thickness proximal and distal gut remnants. Results. Concomitant hGH and EGF treatment up-regulates glucose (100%), glutamine (80%), and leucine (60%) transport in the proximal remnant; alanine (150%) and arginine (400%) transport in the distal remnant; and microvillus height (25% to 35%) both proximally and distally. Serum IGF-I levels and gross villus heights were not different among groups. Conclusions. Co-infusion of hGH and EGF accelerates intestinal adaptation after ME in an additive, nutrient-dependent, and site-specific fashion via enhanced nutrient transport as well as microvillus hypertrophy.

Background: After massive enterectomy, remnant intestine undergoes compensatory adaptation. A combination of human growth hormone (hGH) and a glutamine-enriched modified diet induces further adaptation in patients with short bowel syndrome (SBS) on long-term total parenteral nutrition. The specific actions of each component, however, are not well-defined. Methods: New Zealand White rabbits were randomized to control, sham operation, or SBS (70% midjejunoileal resection) groups and treated with either hGH or saline. Sodium-dependent uptake of glucose, glutamine, alanine, leucine, and arginine into brush border membrane vesicles was quantitated. Serum insulin-like growth factor-I (IGF-I) levels were determined by immunoradiometric assay. Mucosal mRNA expression of IGF-I and IGF binding protein 4 (IGFBP-4) was evaluated by Northern blot analysis using rat cDNA probes. Results: Glutamine and leucine transports were 33 and 39% greater, respectively, in the hGH-treated versus saline-treated SBS group (P < 0.05), supporting induction of system B amino acid transport. This upregulation was due, in part, to an 88% increase in glutamine carrier capacity (V max ) with no change in carrier affinity (K m ). Both hGH treatment and SBS increased serum IGF-I levels without direct correlation with increased nutrient transport. IGFBP-4 mRNA expression in small bowel mucosa of saline-treated SBS animals was significantly greater than saline-treated unoperated control values. Mucosal IGFBP-4 mRNA was not significantly altered from control in the other study groups. IGF-I mRNA expression was not detected in mucosa, but weak hybridization was noted in rabbit liver. Conclusions: Human growth hormone accelerates early adaptation in SBS by upregulation of system B carrier capacity. Serum IGF-I levels and mucosal IGF-I and IGFBP-4 mRNA expression did not directly correlate with this enhanced nutrient transport, suggesting that hGH might exert its adaptive effects by mechanisms that are independent from the IGF system in this model.

Circulating ghrelin, produced primarily in the stomach, is a powerful orexigen. Ghrelin levels are elevated in states of hunger, but rapidly decline postprandially. Early alterations in ghrelin levels in morbidly obese patients undergoing weight reduction surgery may be attributed to gastric partitioning.Thirty-four patients underwent Roux-en-Y gastric bypass with a completely divided gastroplasty to create a 15-mL vertically oriented gastric pouch. Eight other patients underwent other gastric procedures that did not involve complete division of the stomach, including 4 vertical banded gastroplasties and 4 antireflux surgical procedures. Six additional patients undergoing antireflux surgery served as lean control subjects. Plasma samples were obtained before surgery and immediately after surgery. In a substudy, plasma was collected after Roux-en-Y limb formation and after dividing the stomach to identify any changes in plasma ghrelin levels.Tertiary university medical center.Ghrelin levels at different stages of surgical intervention.Mean +/- SEM preoperative and postoperative ghrelin levels in the gastric bypass group were 355 +/- 20 and 246 +/- 13 pg/mL, respectively (P.001). In the vertical banded gastroplasty group and in all patients undergoing antireflux surgery, ghrelin levels were not significantly changed.Compared with morbidly obese humans, lean controls had significantly higher plasma ghrelin levels at baseline. A divided gastroplasty creating a small proximal gastric pouch results in significant early declines in circulating ghrelin levels that are not observed with other gastric procedures. This may explain, in part, the loss of hunger sensation and rapid weight loss observed following gastric bypass surgery.