RESUMEN
BACKGROUND: Dysmotility and postoperative ileus (POI) are frequent major clinical problems post-abdominal surgery. Erythropoietin (EPO) is a multifunctional tissue-protective cytokine that promotes recovery of the intestine in various injury models. While EPO receptors (EPOR) are present in vagal Schwann cells, the role of EPOR in POI recovery is unknown because of the lack of EPOR antagonists or Schwann-cell specific EPOR knockout animals. This study was designed to explore the effect of EPO via EPOR in vagal nerve Schwann cells in a mouse model of POI. RESULTS: The structural features of EPOR and its activation by EPO-mediated dimerization were understood using structural analysis. Later, using the Cre-loxP system, we developed a myelin protein zero (Mpz) promoter-driven knockout mouse model of Schwann cell EPOR (MpzCre-EPORflox/flox / Mpz-EPOR-KO) confirmed using PCR and qRT-PCR techniques. We then measured the intestinal transit time (ITT) at baseline and after induction of POI with and without EPO treatment. Although we have previously shown that EPO accelerates functional recovery in POI in wild type mice, EPO treatment did not improve functional recovery of ITT in POI of Mpz-EPOR-KO mice. CONCLUSIONS: To the best of our knowledge, this is the first pre-clinical study to demonstrate a novel mouse model of EPOR specific knock out on Schwan cells with an effect in the gut. We also showed novel beneficial effects of EPO through vagus nerve Schwann cell-EPOR in intestinal dysmotility. Our findings suggest that EPO-EPOR signaling in the vagus nerve after POI is important for the functional recovery of ITT.
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Eritropoyetina , Receptores de Eritropoyetina , Ratones , Animales , Receptores de Eritropoyetina/metabolismo , Eritropoyetina/metabolismo , Células de Schwann/metabolismo , Transducción de Señal , Ratones Noqueados , Motilidad GastrointestinalRESUMEN
Background: Liver cirrhosis is a relevant comorbidity with increasing prevalence. Postoperative decompensation and development of complications in patients with cirrhosis remains a frequent clinical problem. Surgery has been discussed as a precipitating event for decompensation and complications of cirrhosis, but the underlying pathomechanisms are still obscure. The aim of this study was to analyze the role of abdominal extrahepatic surgery in cirrhosis on portal pressure and fibrosis in a preclinical model. Methods: Compensated liver cirrhosis was induced using tetrachlormethane (CCL4) inhalation and bile duct ligation (BDL) models in rats, non-cirrhotic portal hypertension by partial portal vein ligation (PPVL). Intestinal manipulation (IM) as a model of extrahepatic abdominal surgery was performed. 2 and 7 days after IM, portal pressure was measured in-vivo. Hydroxyproline measurements, Sirius Red staining and qPCR measurements of the liver were performed for evaluation of fibrosis development and hepatic inflammation. Laboratory parameters of liver function in serum were analyzed. Results: Portal pressure was significantly elevated 2 and 7 days after IM in both models of cirrhosis. In the non-cirrhotic model the trend was the same, while not statistically significant. In both cirrhotic models, IM shows strong effects of decompensation, with significant weight loss, elevation of liver enzymes and hypoalbuminemia. 7 days after IM in the BDL group, Sirius red staining and hydroxyproline levels showed significant progression of fibrosis and significantly elevated mRNA levels of hepatic inflammation compared to the respective control group. A progression of fibrosis was not observed in the CCL4 model. Conclusion: In animal models of cirrhosis with continuous liver injury (BDL), IM increases portal pressure, and development of fibrosis. Perioperative portal pressure and hence inflammation processes may be therapeutic targets to prevent post-operative decompensation in cirrhosis.
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Inflammation theory has suggested that the pathogenesis of postoperative ileus (POI) involves the steroid receptor coactivator-3 (SRC-3). Therefore, we investigated the role of SRC-3 in the muscles of the small intestine using a mouse POI model. Here, we reported that intestinal manipulation (IM) significantly reduced the extent of phenol red migration in the entire gastrointestinal tract, and the calculated geometric center (GC) value in wild-type (WT) mice at 24 h after surgery was higher than that in the knockout (KO) mice and in the sham-operated control group. The expression of SRC-3 was upregulated in the mouse intestinal muscularis at 24 h after surgical manipulation, and the mRNA and protein levels of inflammatory cytokines were upregulated compared with those in the control group. At 24 h after IM, the number of neutrophils in the experimental group was significantly higher than that in the control group; in the IM group, the number of neutrophils in the SRC-3-/- mice was markedly higher than that in the WT mice. At 24 h after IM, the myeloperoxidase (MPO) activity in the experimental group was significantly higher than that in the control group. In the IM group, the MPO activity of the SRC-3-/- mice was markedly higher than that of the WT mice. In summary, proinflammatory cytokines, the number of neutrophils, and the MPO activity were significantly increased in the muscularis of the jejunum and ileum of KO mice after IM compared with those of the WT mice, indicating that SRC-3 might play a protective role in POI.
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Citocinas/metabolismo , Motilidad Gastrointestinal , Ileus/metabolismo , Mediadores de Inflamación/metabolismo , Intestino Delgado/metabolismo , Músculo Liso/metabolismo , Coactivador 3 de Receptor Nuclear/metabolismo , Complicaciones Posoperatorias/metabolismo , Animales , Modelos Animales de Enfermedad , Femenino , Ileus/etiología , Ileus/inmunología , Ileus/fisiopatología , Intestino Delgado/inmunología , Intestino Delgado/fisiopatología , Yeyuno/inmunología , Yeyuno/metabolismo , Yeyuno/fisiopatología , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Músculo Liso/inmunología , Músculo Liso/fisiopatología , Infiltración Neutrófila , Coactivador 3 de Receptor Nuclear/genética , Peroxidasa/metabolismo , Complicaciones Posoperatorias/etiología , Complicaciones Posoperatorias/inmunología , Complicaciones Posoperatorias/fisiopatología , Técnicas de Cultivo de TejidosRESUMEN
BACKGROUND: Dysmotility and postoperative ileus (POI) are major clinical problems after surgical trauma and it is associated with increased intestinal inflammation and oxidative stress. Despite the high occurrence of POI following intra-abdominal surgeries, no effective treatment is currently available. Erythropoietin (EPO) is a multifunctional tissue-protective cytokine with potent anti-inflammatory and anti-oxidative properties, and it is an FDA approved medicine for clinical use. While both EPO and EPO receptors (EPOR) are widely expressed in the gut, the role of EPO in POI is largely unknown. This study was designed to explore the possible beneficial effect of EPO in a mouse model of POI. METHODS: Mice were subjected to intestinal manipulation to induce standard POI and intestinal transit time was determined at 24-h post-injury with or without EPO treatment (5000 units/kg, once, IP, immediately after intestinal trauma). Intestinal samples were harvested for histological and immunohistochemical analysis. RESULTS: Systemic EPO significantly improved intestinal transit time compared with control group and it was associated with significantly increased levels of tissue macrophages and reduced levels of oxidative stress. CONCLUSIONS AND INFERENCES: This is the first pre-clinical study to document novel beneficial effects of EPO in gut dysmotility and our findings suggest that the beneficial effects of EPO in POI is predominantly mediated by its anti-oxidative and immunomodulatory properties.
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Eritropoyetina/farmacología , Motilidad Gastrointestinal/efectos de los fármacos , Seudoobstrucción Intestinal , Recuperación de la Función/efectos de los fármacos , Animales , Modelos Animales de Enfermedad , Masculino , Ratones , Ratones Endogámicos C57BL , Complicaciones PosoperatoriasRESUMEN
BACKGROUND: Local release of mast cell proteases during gastrointestinal surgery is associated with the inhibition of motility and postoperative ileus (POI). We determined whether activation of intramuscular mast cell affects the motor patterns of the human ileum and colon and whether proteases are involved. METHODS: Motor response of ileal and colonic circular muscle strips was measured in organ bath. Mast cell degranulation was induced by compound 48/80 (c48/80; 25-675 µg/mL). Motor response was quantified as tone, rhythmic phasic contractions (RPCs) and contractions to electric field stimulation (EFS; 40 Hz), and bethanechol-evoked contractions. Ketotifen (10(-6) mol/L) and a protease inhibitor cocktail (P8340) were used to evaluate the role of mast cell mediators. KEY RESULTS: (a) c48/80 impaired the spontaneous and the electrically evoked motor response in small bowel and colonic strips (sigmoid colon EC50 : 460.0 µg/mL for RPCs and 8.9 µg/mL for electrically evoked contraction amplitudes) and bethanechol-evoked contractions. (b) Preincubation with ketotifen (10(-6) mol/L, 1 h) prevented the impairment of RPCs and EFS-evoked contractions in the sigmoid colon and ileum but not in the right colon. (c) Preincubation with P8340 also prevented the impairment of contractions in the sigmoid colon but not in the ileum or the right colon. CONCLUSIONS & INFERENCES: Mast cell degranulation by c48/80 inhibits the spontaneous and the nerve-mediated motor response in the human ileum and colon. The effect is partially mediated by mast cell proteases and could be relevant in the pathophysiology of POI.
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Degranulación de la Célula , Colon Sigmoide/fisiología , Íleon/fisiología , Mastocitos/fisiología , p-Metoxi-N-metilfenetilamina/farmacología , Adulto , Anciano , Anciano de 80 o más Años , Colon Sigmoide/efectos de los fármacos , Femenino , Humanos , Íleon/efectos de los fármacos , Ileus/complicaciones , Ileus/fisiopatología , Técnicas In Vitro , Cetotifen/farmacología , Masculino , Mastocitos/efectos de los fármacos , Persona de Mediana Edad , Contracción Muscular/efectos de los fármacos , Complicaciones Posoperatorias , Inhibidores de Proteasas/farmacologíaRESUMEN
BACKGROUND: Postoperative ileus (POI) is characterized by impaired gastrointestinal motility resulting from intestinal handling-associated inflammation. The introduction of laparoscopic surgery has dramatically reduced the duration of POI. However, it remains unclear to what extent this results in a reduction of intestinal inflammation. The aim of the present study is to compare the degree of intestinal inflammation and gastrointestinal transit following laparoscopic surgery and open abdominal surgery. METHODS: Mice were subjected to laparoscopic surgery or laparotomy alone or, in combination with standardized intestinal manipulation of the small bowel (IM). Gastrointestinal transit and intestinal inflammation were assessed 24 h after surgery by the number of myeloperoxidase (MPO) positive cells and the level of cytokine expression. The recovery time and the degree of inflammation were also analyzed in patients subjected to colectomy under open conditions (laparotomy) or laparoscopic conditions. KEY RESULTS: Mice undergoing IM by laparotomy (open IM), but not by laparoscopy (Lap IM) developed a significant delay in gastrointestinal transit compared to laparotomy or laparoscopy alone. In addition, there was significant intestinal inflammation only after open IM. Similarly, cytokine levels in peritoneal lavage fluid were lower while recovery time was faster in patients subjected to colectomy under laparoscopic conditions compared to open colectomy. CONCLUSIONS & INFERENCES: Our data confirms that intestinal inflammation is underlying the delayed gastrointestinal transit observed after open surgery. Most importantly, we demonstrate that intestinal inflammation under laparoscopic conditions is significantly lower compared to open surgery, most likely explaining the faster recovery following laparoscopic surgery.
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Enteritis/etiología , Ileus/etiología , Enfermedades del Yeyuno/etiología , Laparoscopía/efectos adversos , Animales , Modelos Animales de Enfermedad , Enteritis/metabolismo , Femenino , Tránsito Gastrointestinal , Humanos , Interleucinas/metabolismo , Enfermedades del Yeyuno/metabolismo , Ratones , Ratones Endogámicos C57BLRESUMEN
BACKGROUND & AIMS: Postoperative ileus (POI) is a common consequence of abdominal surgery that increases the risk of postoperative complications and morbidity. We investigated the cellular mechanisms and immune responses involved in the pathogenesis of POI. METHODS: We studied a mouse model of POI in which intestinal manipulation leads to inflammation of the muscularis externa and disrupts motility. We used C57BL/6 (control) mice as well as mice deficient in Toll-like receptors (TLRs) and cytokine signaling components (TLR-2(-/-), TLR-4(-/-), TLR-2/4(-/-), MyD88(-/-), MyD88/TLR adaptor molecule 1(-/-), interleukin-1 receptor [IL-1R1](-/-), and interleukin (IL)-18(-/-) mice). Bone marrow transplantation experiments were performed to determine which cytokine receptors and cell types are involved in the pathogenesis of POI. RESULTS: Development of POI did not require TLRs 2, 4, or 9 or MyD88/TLR adaptor molecule 2 but did require MyD88, indicating a role for IL-1R1. IL-1R1(-/-) mice did not develop POI; however, mice deficient in IL-18, which also signals via MyD88, developed POI. Mice given injections of an IL-1 receptor antagonist (anakinra) or antibodies to deplete IL-1α and IL-1ß before intestinal manipulation were protected from POI. Induction of POI activated the inflammasome in muscularis externa tissues of C57BL6 mice, and IL-1α and IL-1ß were released in ex vivo organ bath cultures. In bone marrow transplantation experiments, the development of POI required activation of IL-1 receptor in nonhematopoietic cells. IL-1R1 was expressed by enteric glial cells in the myenteric plexus layer, and cultured primary enteric glia cells expressed IL-6 and the chemokine monocyte chemotactic protein 1 in response to IL-1ß stimulation. Immunohistochemical analysis of human small bowel tissue samples confirmed expression of IL-1R1 in the ganglia of the myenteric plexus. CONCLUSIONS: IL-1 signaling, via IL-1R1 and MyD88, is required for development of POI after intestinal manipulation in mice. Agents that interfere with the IL-1 signaling pathway are likely to be effective in the treatment of POI.