The tankyrase inhibitor G007-LK inhibits small intestine LGR5+ stem cell proliferation without altering tissue morphology

Abstract Background The WNT pathway regulates intestinal stem cells and is frequently disrupted in intestinal adenomas. The pathway contains several potential biotargets for interference, including the poly-ADP ribosyltransferase enzymes tankyrase1 and 2. LGR5 is a known WNT pathway target gene and marker of intestinal stem cells. The LGR5+ stem cells are located in the crypt base and capable of regenerating all intestinal epithelial cell lineages. Results We treated Lgr5-EGFP-Ires-CreERT2;R26R-Confetti mice with the tankyrase inhibitor G007-LK for up to 3 weeks to assess the effect on duodenal stem cell homeostasis and on the integrity of intestinal epithelium. At the administered doses, G007-LK treatment inhibited WNT signalling in LGR5+ stem cells and reduced the number and distribution of cells traced from duodenal LGR5+ stem cells. However, the gross morphology of the duodenum remained unaltered and G007-LK-treated mice showed no signs of weight loss or any other visible morphological changes. The inhibitory effect on LGR5+ stem cell proliferation was reversible. Conclusion We show that the tankyrase inhibitor G007-LK is well tolerated by the mice, although proliferation of the LGR5+ intestinal stem cells was inhibited. Our observations suggest the presence of a tankyrase inhibitor-resistant cell population in the duodenum, able to rescue tissue integrity in the presence of G007-LK-mediated inhibition of the WNT signalling dependent LGR5+ intestinal epithelial stem cells.

Carcinogenesis of the upper gastrointestinal tract induced by N-methyl-N'-nitro-nitrosoguanidine and reflux of duodenal contents in the rat / Carcinogênese do trato gastrointestinal alto induzida pelo N-methyl-N'-nitro-nitrosoguanidina e pelo refluxo duodenogástrico no rato

PURPOSE: To investigate the combined effects of reflux of duodenal contents through the pylorus and treatment with N-methyl-N'-nitro-nitrosoguanidine (MNNG) on the development of lesions in the glandular stomach, at the gastrojejunal anastomosis and in the forestomach of rats. METHODS: Eighty Male Wistar rats were divided into 4 groups: G1: MNNG + Reflux, G2: Reflux, G3: MNNG and G4: Gastrostomy. MNNG was given in the drinking water (100 mg/ml) for 12 weeks and then two groups (G1 and G2) were submitted to a gastrojejunal anastomosis followed by section of the afferent loop and suture of both stumps to allow reflux of duodenal contents through the pylorus. The animals were sacrificed 18 and 36 weeks after surgery. The lesions obtained in the antral mucosa, at the gastrojejunal anastomosis and in the forestomach were analysed histologically. RESULTS: Duodenal reflux induced proliferative lesions at both glandular and squamous mucosa of the stomach. In the antrum, adenomatous hyperplasia (AH) was observed in 20% and 50% of the animals at the 18th and 36th weeks respectively. Aditionally 85% of the animals presented AH at the gastrojejunal anastomosis and 60% developed squamous hyperplasia at the squamous portion of the stomach. MNNG treatment plus duodenal reflux enhanced the development of malignant tumors at both glandular and squamous mucosa, since there were 30% of antral adenocarcinomas and 45% of squamous carcinomas at the 18th week and the frequency of these malignant tumors rose to 50% in the antrum and 65% in the squamous mucosa at the 36th week. CONCLUSION: The reflux of duodenal contents through the pylorus enhanced the development of proliferative lesions, benign and malignant, in the glandular stomach and in the forestomach of rats.

OBJETIVO: Investigar os efeitos do refluxo duodenogástrico e sua interação com o cancerígeno químico N-methil-N'-nitro-nitrosoguanidina (MNNG) no desenvolvimento de lesões no estômago glandular, anastomose gastrojejunal e no estômago escamoso do rato. MÉTODOS: Foram utilizados 80 ratos Wistar divididos em 4 grupos: G1: MNNG + Refluxo, G2: Refluxo, G3: MNNG e G 4: Gastrostomia. O MNNG foi oferecido na água de beber (100mg/ml) por 12 semanas. A seguir foi feita anastomose gastrojejunal na porção glandular do estômago nos grupos G1 e G2, com secção da alça aferente junto ao estômago e sutura de ambos os cotos para permitir o refluxo do conteúdo duodenal para o estômago pelo piloro. Os animais foram sacrificados 18 e 36 semanas após a cirurgia. As lesões identificadas foram submetidas à exame histopatológico. RESULTADOS: O refluxo duodenogástrico levou ao desenvolvimento de lesões proliferativas no estômago glandular e na porção escamosa. No antro, hiperplasia adenomatosa (HA) foi diagnosticada em 20 e 50% dos animais (G2) na 18ª e 36ª semanas, respectivamente. Na anastomose gastrojejunal 85 por cento dos animais (G2) apresentaram HA e 60% apresentaram hiperplasia escamosa no estômago escamoso, na 36ª semana. No grupo MNNG+Refluxo foram identificados na 18ª semana, 30% adenocarcinomas no antro e 45%carcinomas escamosos. A freqüência destas lesões malignas aumentou, respectivamente, para 50% e 65% na 36ª semana. CONCLUSÃO: O refluxo duodenogástrico potencializou o desenvolvimento de lesões proliferativas benignas e malignas no estômago glandular e em sua porção escamosa, no rato.

En bloc Resection for Right Colon Cancer Directly Invading Duodenum or Pancreatic Head

PURPOSE: We undertook this study to analyze clinical features and surgical outcome of en bloc resections of the right side colon cancer directly invading duodenum and/or pancreatic head. MATERIALS AND METHODS: The records of all patients who underwent en bloc resection of duodenum and/or pancreas for right colon cancers were analyzed retrospectively. From September 1994 to September 2006, 1,016 patients underwent curative right hemicolectomy. Nine patients (0.9%) had en bloc resection of a right side colon cancer with duodenum or pancreatic head invasion. RESULTS: The median operative time was 320 minutes (range, 200-420) and the median blood loss was 700 mL (range, 100-2,000). The mean size of tumor was 6.6 cm (range, 3.2-10.7). The mean preoperative carcinoembryonic antigen (CEA) was 10.6 ng/mL (range, 0.2-50.8). There was no 30 day perioperative mortality. The median disease-free survival was 23.5 months [95% confidence interval (CI) 5.2-41.8] and the median overall survival was 28.1 months (95% CI 9.7-46.5). CONCLUSIONS: In patients with locally advanced right side colon cancer that directly invades the duodenum or pancreas can be safely resected with curative potential with minimum morbidity and mortality. Long term disease free survival can occur in a significant number of patients undergoing curative en bloc resection in this particular subset of patients.

Histological study on the effect of thymoquinone -an immunomodulator- on duodenum and superior mesenteric lymph nodes of cortisone treated rats

Thymoquinone [TQ] was demonstrated as the major active component of Nigella sativa seed oil. It was reported that thymoquinone has an anti-inflammatory and immune-stimulatory effects. On the other hand, cortisone is one of the steroid hormones that used as an anti-inflammatory and immune-suppressor drug. In the present study, the effects of both thymoquinone and cortisone were studied on the histological structure of both duodenum and superior mesenteric lymph nodes of rats. It was noticed that there was an increase of the duodenum mucosal mucus-secreting cells and some nuclear changes in the nuclei of the Brunner's gland cells. There was an increase in number of macrophages, plasma cells and lymphocytes in the superior mesenteric lymph nodes. These findings were discussed according the anti-inflammatory and the immunestimulatory effects of thymoquinone as a point of view, and the anti-inflammatory and immune-suppressing effects of cortisone on another point of view. It was concluded that thymoquinone has an anti-inflammatory and immune sthnulatory effects on rat duodenum and superior mesenteric lymph nodes. However, more investigations are still in need to recommend the replacement of this natural ingredient instead of cortisone with its side effects

Activation of neural cholecystokinin-1 receptors induces relaxation of the isolated rat duodenum which is reduced by nitric oxide synthase inhibitors

Cholecystokinin (CCK) influences gastrointestinal motility, by acting on central and peripheral receptors. The aim of the present study was to determine whether CCK has any effect on isolated duodenum longitudinal muscle activity and to characterize the mechanisms involved. Isolated segments of the rat proximal duodenum were mounted for the recording of isometric contractions of longitudinal muscle in the presence of atropine and guanethidine. CCK-8S (EC50: 39; 95 percent CI: 4.1-152 nM) and cerulein (EC50: 58; 95 percent CI: 18-281 nM) induced a concentration-dependent and tetrodotoxin-sensitive relaxation. Nomeganitro-L-arginine (L-NOARG) reduced CCK-8S- and cerulein-induced relaxation (IC50: 5.2; 95 percent CI: 2.5-18 æM) in a concentration-dependent manner. The magnitude of 300 nM CCK-8S-induced relaxation was reduced by 100 æM L-NOARG from 73 ± 5.1 to 19 ± 3.5 percent in an L-arginine but not D-arginine preventable manner. The CCK-1 receptor antagonists proglumide, lorglumide and devazepide, but not the CCK-2 receptor antagonist L-365,260, antagonized CCK-8S-induced relaxation in a concentration-dependent manner. These findings suggest that CCK-8S and cerulein activate intrinsic nitrergic nerves acting on CCK-1 receptors in order to cause relaxation of the rat duodenum longitudinal muscle.

Prokinetic effect of clarithromycin and azithromycin in vitro study on rabbit duodenum

There is a continuous search for new prokinetic agents for use in gastro intestinal hypomotility. Erythromycin, a macrolide, is one of them. In this study we observed effect of other macrolides, i.e. clarithromycin and azithromycin on rabbit duodenum and compared with that of neostigmine. Both these drugs produced significant prokinetic effect with EC[50] 0.4 micro g/ml and 0.29 micro g/ml respectively. Effect of clarithromycin was well sustained as compared to that of azithromycin, so it seemed to be a better prokinetic agent

Brainstem origin of duodenal vagal preganglionic parasympathetic neurons. A WGA-HRP study in the ferret (Mustela Putorius Furo), a human model

The projections of vagal brainstem neurons to the duodenal segment of the gastrointestinal tract were studied in the ferret using the WGA-HRP neurohistochemical technique. Fourteen adult ferrets with weights ranging from 800 gm to 1500 gm were used for the study. The muscular wall of the duodenum of six ferrets was injected with 0.1 ml of 5 WGA-HRP in 0.5 M sodium chloride. The eight remaining ferrets were used as controls. Two of these had injections of 0.1 ml normal saline into the muscular wall of the duodenum. The second set of two ferrets was injected with 0.1 ml of 5 WGA-HRP in buffer after bilateral truncal vagotomy. The third set of two ferrets received intraperitoneal injection of 0.1 ml of 5 WGA-HRP while, in the last set, the tracer was injected into the hepatic portal vein. Following the injections, the ferrets were allowed to survive for 48-72 hours after which each ferret was perfused transcardially first with normal saline followed by a fixative containing 1 paraformaldehyde and 1.25 glutaraldehyde in 0.1 M phosphate buffer, pH 7.4 at room temperature and finally with 10 buffered sucrose at 4 degrees C. Transverse serial frozen sections of the brainstem were then taken and processed for WGA-HRP neurohistochemistry and were analyzed under light and dark-field illuminations. The analyses of the sections taken from the six ferrets injected with WGA-HRP revealed neurons labelled with the tracer in the dorsal motor nucleus of the vagus nerve (DMNV). Sections taken from the control ferrets did not reveal any WGA-HRP labelled neurons in the brainstem

Comportamento motor, sob estímulo farmacológico, de segmentos duodenais pré e pós-papilar de ratos albinos / Motor reaction, under the pharmacological stimulus of the isolated duodenal pre-papillary and post papilary section of albino rats

Cada segmento do tubo digestivo tem um padrao próprio de motilidade e toda atividade motora resulta fundamentalmente das propriedades elétricas do músculo liso. Em sua atividade motora o duodeno apresenta funçoes de mistura e progressao, como também participa do controle do esvaziamento gástrico. Os movimentos do complexo antro-piloro-bulbar sao considerados ponto de partida para o peristaltismo intestinal, inclusive com movimentos reversos, sendo que encontramos normalmente uma diminuiçao progressiva da freqüência das ondas do duodeno ao íleo terminal. Considerando que o duodeno tem características embriológicas, anatômicas, histológicas e bioquímicas que o destacam na fisiologia digestiva, interessou-nos o comportamento motor do segmento duodenal pré e pós-papilar sob estímulo farmacológico. Na realizaçao do tranbalho foram utilizados dez ratos albinos dos quais foram retirados os segmentos duodenais pré e pós-papilar. Estes foram divididos em: grupos I e II, pré e pós-papilar, respectivamente. Em cada segmento foram obtidos as diferenças porcentuais (delta percentual) do número de ondas e da amplitude máxima no movimento espontaneo e sob estimulo farmacológico do cálcio, potássio, acetilcolina e nor-adrenalina. Foram analisados segundo a açao das drogas dentro de cada grupo e, também, analisadas a açao de cada droga, comparadas entre os grupos. Os resultados mostraram diferenças no D percentual do número de ondas sob estímulo farmacológico quando comparados ao movimento espontâneo. Na amplitude máxima, porém, nao houve diferença significante. Nas condiçoes de realizaçao do trabalho, pode-se concluir entao, que o comportamento motor sob estímulo farmacológico do segmento duodenal pré-papilar isolado de rato é diferente do pós-papilar.

Gastroduodenal resistance and neural mechanisms involved in saline flow decrease elicited by acute blood volume expansion in anesthetized rats

We have previously demonstrated that blood volume (BV) expansion decreases saline flow through the gastroduodenal (GD) segment in anesthetized rats (Xavier-Neto J, dos Santos AA & Rola FH (1990) Gut, 31: 1006-1010). The present study attempts to identify the site(s) of resistance and neural mechanisms involved in this phenomenon. Male Wistar rats (N = 97,200-300 g) were surgically manipulated to create four gut circuits: GD, gastric, pyloric and duodenal. These circuits were perfused under barostatically controlled pressure (4 cmH2O). Steadysate changes in flow were taken to reflect modifications in circuit resistances during three periods of time: normovolemic control (20 min), expansion (10-15 min), and expanded (30 min). Perfusion flow rates did not change in normovolemic control animals over a period of 60 min. BV expansion (Ringer bicarbonate, 1 ml/min up to 5 percent body weight) significantly (p<0.05) reduced perfusion flow in the GD (10.3 + 0.5 to 7.6 + 0.6 ml/min), pyloric (9.0 + 0.6 to 5.6 + 1.2 ml/min) and duodenal (10.8 + 0.4 to 9.0 + 0.6 ml/min) circuits, but not in the gastric circuit (11.9 + 0.4 to 10.4 + 0.6 ml/min). Prazosin (1 mg/kg) and yohimbine (3 mg/kg) prevented the expansion effect on the duodenal but not on the pyloric circuit. Bilateral cervical vagotomy prevented the expansion effect on the pylorus during the expansion but not during the expanded period and had no effect on the duodenum. Atropine (0.5 mg/kg), hexamethonium (10 mg/kg) and propranolol (2 mg/kg) were ineffective on both circuits. These results indicate that 1) BV expansion increases the GD resistance to liquid flow, 2) pylorus and duodenum are important sites of resistance, and 3) yohimbine and prazosin prevented the increase in duodenal resistance and vagotomy prevented it partially in the pylorus.

Evidence for the participation of the L-arginine-nitric oxide pathway in neurally induced relaxation of the isolated rat duodenum

1. Electrical field stimulation (EFS) of intrinsic nerves in the rat proximal duodenum induces a frequency-dependent non-adrenergic-non-cholinergic (NANC) relaxation response. 2. The inhibitors of L-arginine-NO synthase L-NG-nitro arginine methyl-ester (L-NAME) and L-NOARG (L-NG-nitro arginine) reduced the NANC relaxations elicited by EFS in a dose- and time-dependent manner; L-NOARG was two times more potent than L-NAME (IC50 = 14.3 vs 25.2 microM) and these effects were partially reverted by the addition of 300-1000 microM L-arginine but not of 300-1000 microMD-arginine. Relaxation caused by vasoactive intestinal peptide (VIP; 0.1 microM) or ATP (20 microM) was not blocked by L-NAME or L-NOARG. 3. The magnitude of the blockade caused by L-NAME and L-NOARG was dependent on the frequency of stimulation. At low frequencies (below 1 Hz) both L-NAME and L-NOARG abolished the relaxations, while at 2 to 8 Hz only partial inhibition was observed (maximal inhibition: 44.6 per cent +/- 5.2 and 63.4 per cent +/- 3.4, respectively) 4. The basal tonus of the duodenum was increased by 10-300 microM L-NAME and 10-300 microM L-NOARG and this effect was blocked by 1 mM L-arginine. 5. Nitric oxide generated from acidified NaNO2 caused a dose-dependent (EC50 = 2.75 microM) relaxation of the duodenum which was not affected by 100 microM L-NAME, 100 microM L-NOARG or 1 microM tetrodotoxin (TTX). 6. NADPH-diaphorase positive neurons and fibers identified by histochemistry were present in the myenteric plexus and along both circular and longitudinal muscle fibers indicating that nitric oxide could be synthetized by these neural structures

Determinação do efeito das drogas antiinflamatórias não-esteróides: indometacina, butazona, clinoril, naprosyn, benflogin e inflaril na atividade mitótica da células da medula óssea e das células epiteliais do duodeno / Evaluation of the effect of the non-steroid antiinflammatory drugs: indomethacin, phenylbutazone, sulindac, naproxen, benzydamine and influmic acid on the mitotic activity of the bone marrow cells and on the duodenal epithelial cells

Estudou-se o papel dos antiinflamatórios não esteróides na cinética da proliferação celular das células hematogênicas da medula óssea e das células da mucosa duodenal. A atividade mitótica das células foi avaliada em ratos após 3, 18 e 36 dias de tratamento ininterrupto com as seguintes drogas: Indometacina, Butazona, Clinoril, Naprosyn, Benflogin e Inflaril. A atividade proliferativa celular nesses tecidos foi determinada pela incorporação de timidina triciada (Thymidine-Methyl 3h com atividade específica de 6,7 Ci/mmol) injetada em cada animal por via subcutânea numa única dose 0,25µ ti/g de peso corporal. De um modo geral, as células hematogênicas da medula óssea e as células epiteliais do duodeno mostraram-se sensíveis aos efeitos antimitóticos dos antiinflamatórios não-esteróides testados no presente trabalho

Influence of calcium removal from bath solution on isolated rat duodenum contractions.

In this study whether extracellular Ca++ is essential to produce an increase of tension of isolated rat duodenum by ACh, 5-HT, AVP and KCl-, was examined. KCl- and AVP-evoked contractions were almost totally prevented by Ca++ removal from bath solution. The increase of tension of isolated duodenum caused by ACh or 5-HT was totally prevented after adding nifedipine, a Ca++ channel blocker, into Ca free solution. Our results suggest that ACh or 5-HT utilizes intracellular as well as extracellular sources of Ca++ to produce contraction in rat duodenum, whereas AVP-or KCl evoked contraction was mainly due to influx of Ca from extracellular sources.

Estudo dos movimentos espontâneos e sob efeitos farmacológicos de segmentos isolados da junçäo duodeno-jejunal de rato / Spontaneous and drug-induced movements of isolated segments of the duodenojejunal junction of rat

Estudou-se, "in vitro", a motilidade espontânea e sob efeito farmacológico da junçäo duodenojejunal de ratos. Tomou-se três segmentos: pré, no nível e após a junçäo. Os registros foram feitos em quimógrafo com papel enfumaçado. A acetilcolina e fisostigmina diminuíram as amplitudes de contraçöes nos três segmentos, bem como a noradrenalina e procaína. A freqüência das contraçöes espontâneas apresentou um gradiente decrescente no sentido crânio-caudal, observado tambem sob o efeito da noradrenalina. Sob o efeito acetilcolina e fisostigmina elas se igualaram. Näo houve diferença quanto ao tipo de ondas (I, II e III), sendo mais frequüente a onda I. Elas näo se alteraram sob a açäo da acetilcolina e noradrenalina, e a fisostigmina alterou as características das ondas. O efeito da procaína sobre as ondas foi marcante, diferenciando, nitidamente, o segmento pré da transiçäo duodenojejunal. O tono foi igual nos três segmentos. A acetilcolina diferenciou o segmento pré do pós-junçäo duodenopejunal. A noradrenalina e a procaina diminuiram o tono e a fisostigmina aumentou-o igualmente nos três segmentos

Blockade of the bradykinin-evoked diphasic response of isolated rat duodenum by the crude extract of and compounds obtained from Mandevilla velutina

The influence of an aqueous/ethanolic extract of M. velutina (CE) and of two compound (MV 8608 and MV 8612) purified from the plant on BK-, Ad- and K+ -induced responses of the rat duodenum was analyzed in vitro. In preparations precontracted with Ca2+, the CE (1 and 2 mg/ml) markedly inhibited BK -induced relaxation in a dose-dependent manner but was less effective against Ad-induced relaxation. The CE (0.5 mg/ml) also antagonized BK -induced relaxation and contraction of strips bathed in normal medium. The two compounds from M. velutina (10 and 20 microng/ml) also promoted a dose-dependent blockade of both responses to BK, only slightly depressing the response to K+

Açäo da amiodarona sobre as contraçöes induzidas por acetilcolina em duodeno isolado de rato / Effect of amiodarone on contractions induced by acetylcholine in isolated rat duodenum

O estudo dos efeitos da amiodarona sobre as respostas contráteis de duodeno isolado de rato induzidas por acetilcolina (5x10 -8M a 5x10 4-M), mostram que as contraçöes foram antagonizadas da maneira dose-dependente por amiodarona em concentraçöes de 2,5x10 -5M. A inibiçäo foi caracterizada como antagonismo näo-competitivo, pois houve reduçäo progressiva da resposta máxima. O valor médio de pD'2 com seu erro padräo foi calculado em 3,92 + ou - 0,31