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sT1765

GLP-2 Stimulates Mucosal Microcirculation Measured by Laser Doppler Flowin Jejunal Stoma of Patients With Short Bowel SyndromePalle B. Jeppesen, Per M. Hellström

Objective: Glucagon-like peptide-2 (GLP-2) is an endogenous meal-stimulated hormonesecreted by L-cells of terminal ileum and colon. In clinical trials with GLP-2 in short bowelsyndrome (SHS) we noted increased redness and size of stoma nipples. GLP-2 receptors areco-localized with endothelial nitric oxide synthase (eNOS) and prerequisites for blood floweffects of GLP-2 are fulfilled. We used native GLP-2 in combination with saline and to studylaser Doppler flow (LDF) in jejunal stomas in man. Methods: 5 patients with SBS; smallbowel <150 cm ending in a jejunostomy were studied. Stoma nipple blood perfusion wasmeasured circumferentially with a designed LDF probe (Periflux 4001, wavelength 780nm,time constant 0.2s, sampling 32Hz). All patients were studied in fasting state and supineposition on 4 occasions; each period lasting 120 min. First no treatment was given, second1 L saline was infused IV, third 5mL of adrenalin 0.1mg/mL was instilled into the stoma,and fourth 1mL of GLP-2 800μg/mL SC. Results were statistically analyzed by two-wayANOVA. Values are mean±SEM. Results: GLP-2 caused a marked increase of LDF, whileother groups (no treatment, saline or adrenaline) displayed no change in LDF. The effectwas long-lasting and persisted significantly elevated over 105 min. The AUC over the wholestudy period was greatly increased in the GLP-2 group with 56219±4628 PU min (p<0.001)as compared to 32283±6120 in untreated, 32531±6109 with saline and 32166±6306 withadrenaline. Conclusions: GLP-2 at clinical dosage increases intestinal microcirculation, whichdoubled during 2 hours after dosing. We suggest that GLP-2 promotes cellular growthinvolving increased microcirculation as prerequisite for trophic effects in the gut.

Laser Doppler flow shows doubling of mucosal microcirculation in the jejunum of patientsunder GLP-2 stimulation for mucosal growth.

T1766

Transient Gastroparesis and Reduced Ghrelin After Gastric Ischemia andReperfusionSachiko Suzuki, Hidekazu Suzuki, Hitoshi Tsugawa, Juntaro Matsuzaki, Hiroshi Hosoda,Kenji Kangawa, Yoshio Kase, Toshifumi Hibi

Background. The gastrointestinal tract is one of the most susceptible organs to ischemia.We recently reported that post-ischemic alterations of the gastric motility were observedafter gastric ischemia and reperfusion (I/R) alongwith disruption of the interstitial cells of Cajalnetwork and decrease of neuronal nitric oxide synthase-positive neurons (Neurogastroenterol.Motil. 2010, in press). Ghrelin is a gastrointestinal peptide that stimulates growth hormonesecretion and gastrointestinal motility. Ghrelin was demonstrated to attenuate the mucosalinjuries induced by gastric I/R (J. Gastroenterol. Hepatol. 22 :1791, 2007), however, thealterations of the ghrelin dynamics after gastric I/R are not elucidated. The aim of the presentstudy was to investigate the relationship between gastric motility and the ghrelin dynamicsafter gastric I/R. Materials and Methods. 7-week-old male Wistar rats were used after fooddeprivation. Under anesthesia, the celiac artery was clamped for 80 min followed by 12 or48 h reperfusion. The plasma ghrelin level was measured by ELISA. Real-time PCR ofpreproghrelin mRNA and immunohistochemical staining using anti-ghrelin antibody wereperformed after gastric I/R. Results. The plasma acyl-ghrelin levels at 12 h and 48 h afterI/R were significantly lower than those at the corresponding time points in the sham-operatedrats (sham 12 h; 64.5 ± 13.5 fmol/ mL, I/R 12 h; 46.3 ± 9.25 fmol/ mL, sham 48 h; 97.2± 30.3 fmol/ mL, I/R 48 h; 70.9 ± 18.4 fmol/ mL) although the delay in gastric emptyingwas restored at 48 h after I/R. The plasma desacyl-ghrelin levels at 12 h and 48 h after I/R were also lower than those at the corresponding time points in the sham-operated rats(sham 12 h; 834 ± 137 fmol/ mL, I/R 12 h; 663 ± 113 fmol/ mL, sham 48 h; 1092 ± 150fmol/ mL, I/R 48 h; 835 ± 187 fmol/ mL). The expression levels of preproghrelin mRNAwere significantly reduced at 12 h after I/R as compared with the levels at the correspondingtime-point in the sham operates rats, but recovered by 48 h. The count of ghrelin-immunore-active cells was also decreased at 12 h after I/R, but recovered by 48 h. However, the mucosalinjuries persisted throughout the observation period. Conclusion. The consistent decreaseof the plasma ghrelin levels is considered to be attributable to the gastric mucosal injuries.

S-574AGA Abstracts

The decrease of the plasma levels of both acy- and desacyl-ghrelin at 48 h after I/R mightnot have directly affected the gastric motility after I/R, since desacyl-ghrelin induces a delayin gastric emptying, whereas acyl-ghrelin accelerates the rate of gastric emptying.

T1767

Multiple Roles for Protease Activated Receptor 2 (PAR2) Activation in thePromotion of the TH2 ResponseJennifer A. Stiltz, Luigi Notari, Aiping Zhao, Joseph F. Urban, Rex Sun, Terez Shea-Donohue

Introduction: A number of enteric nematodes preferentially infect the upper small intestine(SI) and upregulate Th2 cytokines. The mechanisms involved in the initiation of Th2-mediated immunity remain unclear, but do not appear to require toll-like receptors (TLR).We showed previously that multiple nematode species produce a serine-like protease thatincreases mucosal permeability through activation of PAR2 and availability of PAR2 is criticalfor this effect. Enhanced permeability allows the passage of worm-derived products acrossthe mucosal barrier and facilitates activation of antigen-presenting cells such as dendriticcells (DC). Aim: To determine the contribution of PAR2 to the development of host defenseagainst nematode infection. Methods: To determine the distribution of PAR2 along the GItract, expression was determined by real-time PCR in mucosal cells collected from the upper,middle, and lower SI and from the colon of uninfected BALB/c mice. To investigate theimmune regulation of PAR2, nematode infection-induced changes in jejunal expression weremeasured at 9 days post Nippostrongylus brasiliensis (Nb) infection. PAR2 is required for DCmaturation; therefore, we used JAWS II cells, an immortalized DC line, to assess the effectsof PAR2 activating peptide (SLIGRL, 200nM) on PAR2 expression. Results were comparedto lipopolysaccharide (LPS, 100ng/ml). Results: PAR2 expression was higher than TLR4 in theSI. Nematode infection induced a STAT6-dependent increase in the expression of PAR2(1.0 ±0.1 vs 2.7 ± 0.4 fold) associated with the increase in mucosal permeability. Nematodeinfection upregulated the expression of mucosal mast cell protease-1 (1.0 ± 0.3 vs 125 ±50 fold), a known PAR2 agonist. Treatment of JAWS II cells (Table) with SLIGRL increasedPAR2 expression. LPS alone had no effect on PAR2 expression, but significantly amplifiedthe effects of SLIGRL. Conclusions: These data suggest that 1) preferential expression ofPAR2 vs TLR4 in the SI favors nematode interactions with PAR2; 2) the STAT6-dependentincrease in PAR2 tissue expression during Nb infection serves to maintain PAR2 availabilitythrough a positive feedback mechanism; and 3) infection increases the availability of PAR2agonists that increase permeability and promote DCmaturation by upregulating PAR2 expres-sion on DCs. This effect can be amplified by LPS suggesting a novel interaction betweenTLR4 and PAR2 in DCs.

*p<0.05 v.s. Veh; φ<0.05 v.s. SLIGRL

T1768

Endogenous Antinociceptive Role of Par-4 in Inflammatory but Not in Stress-Induced Colorectal Hyperalgesia in MiceAnita Annahazi, Marta Dabek, Krisztina Gecse, Andras I. Rosztoczy, Richárd Róka, VassiliaTheodorou, Tibor Wittmann, Helene Eutamene, Lionel P. Bueno

Introduction: Activation of proteinase-activated receptor-4 (PAR-4) from the colonic lumenby its agonist AYPGKF-NH2 (PAR-4-AP) or by cathepsin-G (Cat-G), a mediator of theneutrophil granulocytes, has antinociceptive effects to colorectal distension (CRD) in micein basal conditions. Objectives: Our aims were to clarify if PAR-4 activation has antihyperalg-esic effect in two visceral hypersensitivity models, water avoidance stress (WAS) and mildacute 2,4,6-trinitrobenzene sulfonic acid (TNBS) colitis, and to test if endogenous PAR-4activation is present in these conditions. Methods: In anaesthetised C57/BL6 male miceelectrodes were inserted in the abdominal muscle to record abdominal cramps. After recoverymice underwent WAS sessions 1 h/day on 4 consecutive days. A second group of animalswas treated with TNBS (20 mg/kg in 30% ethanol intracolonically, IC). Controls were eitherSHAM-stressed or received 0.9% saline IC. Following the 4th WAS session or 72 h afterthe infusion of TNBS mice were IC infused with 100 μg PAR-4-AP (0.667 mg/mL) or itsvehicle. To test the endogenous role of PAR-4, other groups of TNBS or WAS treated animalsreceived a PAR-4 antagonist (P4pal-10, pepducin, 0.75 mg/kg) or its vehicle intraperitoneally(IP). CRD was performed 1 h after the end of infusion or 30 min after the IP injection byusing a balloon (Fogarthy probe) inflated from 0 to 0.1 mL each steps lasting 10 s with 5min intervals. Results: WAS significantly increased the abdominal EMG response by 760,123, 68 and 22%, at the volumes of 0.02-0.04-0.06-0.08 mL, respectively. PAR-4-APcompletely restored basal sensitivity at all volumes (p<0.05). PAR-4 antagonist did notchange the hyperalgesia provoked by stress (p>0.05). In TNBS colitis, EMG activity waselevated by 1529, 98, 90 and 25% compared to controls for distension volumes from 0.02to 0.08 mL. PAR-4-AP reduced the increase of visceral sensitivity to the baseline for eachvolumes tested (p<0.05). Interestingly, PAR-4 antagonist significantly increased the hyper-sensitivity by 78%, 42% and 19% at the volumes of 0.04, 0.06 and 0.08 mL compared toTNBS alone (p<0.05). Cat-G was elevated in feces after TNBS (17.0 ± 2.6 vs 8.8 ± 1.9 U/mg, p<0.05), but not after WAS (6.9 ± 2.5 U/mg) compared to controls. Conclusions: PAR-4 activation displays analgesic properties in both inflammatory (TNBS) and non-inflammatory(WAS) colorectal hyperalgesia. Further, endogenous PAR-4 activation is present and playsan antinociceptive role in inflammatory, but not in stress induced colorectal hyperalgesia.These data also suggest that Cat-G released by neutrophils in TNBS colitis may be responsiblefor such endogenous activation.