Treatment with the 5-Lipoxygenase Antagonist Zileuton Protects Mice from Postoperative Ileus.

INTRODUCTION: Previous work of our group showed that lipoxygenase (LOX) pathways become activated upon surgical manipulation of the bowel wall and revealed a beneficial immune modulating role of the LOX-derived anti-inflammatory mediator protectin DX in postoperative ileus (POI). While we found a particular role of 12/15-LOX in the anti-inflammatory LOX action during POI, the role of 5-LOX, which produces the pro-inflammatory leukotriene B4 (LTB4), remained unknown. The purpose of this study was to investigate the role of 5-LOX within the pathogenesis of POI in a mouse model. METHODS: POI was induced by intestinal manipulation (IM) of the small bowel in C57BL/6, 5-LOX-/-, and CX3CR1GFP/+. Mice were either treated with a vehicle or with the synthetic 5-LOX antagonist zileuton or were left untreated. Cellular localization of 5-LOX and LTB4 release were visualized by immunofluorescence or ELISA, respectively. POI severity was quantified by gastrointestinal transit (GIT) and leukocyte extravasation into the muscularis externa (ME) by immunohistochemistry. RESULTS: 5-LOX expression was detected 24 h after IM within infiltrating leukocytes in the ME. LTB4 levels increased during POI in wild type but not in 5-LOX-/- after IM. POI was ameliorated in 5-LOX-/- as shown by decreased leukocyte numbers and normalized GIT. Zileuton normalized the postoperative GIT and reduced the numbers of infiltrating leukocytes into the ME. DISCUSSION/CONCLUSION: Our data demonstrate that 5-LOX and its metabolite LTB4 play a crucial role in POI. Genetic deficiency of 5-LOX and pharmacological antagonism by zileuton protected mice from POI. 5-LOX antagonism might be a promising target for prevention of POI in surgical patients.

Author Correction: AIM2 inflammasome-derived IL-1ß induces postoperative ileus in mice.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

AIM2 inflammasome-derived IL-1ß induces postoperative ileus in mice.

Postoperative ileus (POI) is an intestinal dysmotility frequently occurring after abdominal surgery. An orchestrated neuroimmune response within the muscularis externa (ME) involves activation of resident macrophages, enteric glia and infiltration of blood-derived leukocytes. Interleukin-1 receptor type-I (IL1R1) signalling on enteric glia has been shown to be involved in POI development. Herein we investigated the distinct role of the IL1R1 ligands interleukin (IL) -1α and IL-1ß and focused on the mechanism of IL-1ß production. IL-1α and IL-1ß deficient mice were protected from POI. Bone-marrow transplantation studies indicated that IL-1α originated from radio-resistant cells while IL-1ß was released from the radio-sensitive infiltrating leukocytes. Mouse strains deficient in inflammasome formation identified the absent in melanoma 2 (AIM2) inflammasome to be crucial for IL-1ß production in POI. Mechanistically, antibiotic-treated mice revealed a prominent role of the microbiome in IL-1ß production. Our study provides new insights into distinct roles of IL-1α and IL-1ß signalling during POI. While IL-1α release is most likely an immediate passive response to the surgical trauma, IL-1ß production depends on AIM2 inflammasome formation and the microbiome. Selective interaction in this pathway might be a promising target to prevent POI in surgical patients.

Role of Interferon (IFN)α in "Cocktails" for the Generation of (Leukemia-derived) Dendritic Cells (DCleu) From Blasts in Blood From Patients (pts) With Acute Myeloid Leukemia (AML) and the Induction of Antileukemic Reactions.

Strategies to stabilize remissions by specific elimination of residual acute myeloid leukemia (AML) blasts are needed. Leukemia-derived dendritic cell (DCleu/DC) generated from myeloid blasts improve antileukemic T-cell reactivity and install T-cell memory. Interferon (IFN)α-DC methods produce DCleu from chronic myeloid leukemia-patients (pts') blood. Various INFα-containing versus other DC methods were studied to produce DCleu (evaluated by flowcytometry) from AML-pts' blast-containing mononuclear (MNC) or whole blood (WB). After DCleu/DC stimulation in mixed lymphocyte cultures, T cells' potential to gain antileukemic cytotoxicity was studied and correlated with different DC methods and DCleu/DC counts. (1) Generation of DCleu/DC: (a) "IFN-GIT" [containing granulocyte macrophage-colony stimulating factor (GM-CSF)+IFNα+ tumor necrosis factor (TNF)-α] produced DC successfully (≥10% DC, ≥5% DCleu/cells) from AML-MNC (WB) in 54 (56%), "MCM-Mimic" in 76 (75%), "Picibanil" in 83 (64%), and "Calcium-ionophore" in 42 (67%) of cases. Proportions of DC subtypes in MNC (WB) were comparable with all DC methods, (b) IFNα combinations containing only GM-CSF+IFNα or only IFNα showed low efficiency to produce DCleu/DC from MNC (WB) compared with "IFN-GIT." (2) Antileukemic functionality: DCleu/DC-stimulated T cells showed improved leukemia cytotoxicity compared with blast cells or unstimulated T cells. The highest blast proliferation (=insufficient T cells) was seen with "IFN-GIT" DC-stimulated T cells. Probability to respond to immunotherapy or to obtain blast lysis of DC-stimulated T cells correlated with high proportions of DCleu/DC after DC culture, independent of DC-generating methods. (3) Cytokine release profiles: levels of interleukin-6, IFN-γ, and interleukin-2 were significantly lower in DC culture supernatants (from MNC/WB) with "IFN-GIT" compared with "MCM," "Pici," and "Ca" DC supernatants. Our data show that (1) WB culture simulates AML-pts' in vivo situation, (2) DC generation is possible from AML-MNC (WB) with IFNα-containing and other DC methods, (3) successful IFNα-DC generation needs GM-CSF+IFNα+TNF-α (IFN-GIT); however, "IFN-GIT" produces less DCleu/DC compared with other (non-IFNα) DC methods, (4) T cells stimulated with "IFN-GIT"-produced DCleu/DC yielded comparable antileukemic cytotoxicity; however, in cases without achieved blast lysis, an increased blast proliferation was observed.

A comparative study about the immunomodulatory effects of tramadol and metamizole in a murine model of postoperative ileus.

Postoperative ileus (POI) is a common complication after abdominal surgery characterized by motility disturbances leading to increased morbidity and mortality in surgical patients. Intestinal manipulation of the murine small bowel is an established animal model resulting in an increased postsurgical inflammation within the intestinal muscular externa and a delayed gastrointestinal transit. Some analgesics have been shown to affect inflammation. In this study, we compared the immunomodulatory effects of two different analgesics. Mice were treated with tramadol, metamizole or saline as a control in our established POI model. The postoperative inflammatory response was assessed by gene expression of pro-inflammatory cytokines at different time points and immunocytes extravasation into the muscularis externa. Functional motility analyses were performed by a gastrointestinal transit measurement. Metamizole application reduced the pro-inflammatory response after surgery and improved gastrointestinal motility, while tramadol showed no alteration in cytokine gene expression, influx of immunocytes and gastrointestinal transit compared with the controls. In conclusion. we suggest tramadol as analgesia in immunological studies on POI in mice as it does not affect the underlying inflammation of POI.

Leukocyte-Derived Interleukin-10 Aggravates Postoperative Ileus.

Objective: Postoperative ileus (POI) is an inflammation-mediated complication of abdominal surgery, characterized by intestinal dysmotility and leukocyte infiltration into the muscularis externa (ME). Previous studies indicated that interleukin (IL)-10 is crucial for the resolution of a variety of inflammation-driven diseases. Herein, we investigated how IL-10 affects the postoperative ME inflammation and found an unforeseen role of IL-10 in POI. Design: POI was induced by a standardized intestinal manipulation (IM) in C57BL/6 and multiple transgenic mouse strain including C-C motif chemokine receptor 2-/-, IL-10-/-, and LysMcre/IL-10fl/fl mice. Leukocyte infiltration, gene and protein expression of cytokines, chemokines, and macrophage differentiation markers as well as intestinal motility were analyzed. IL-10 serum levels in surgical patients were determined by ELISA. Results: IL-10 serum levels were increased in patient after abdominal surgery. In mice, a complete or leucocyte-restricted IL-10 deficiency ameliorated POI and reduced the postoperative ME neutrophil infiltration. Infiltrating monocytes were identified as main IL-10 producers and undergo IL-10-dependent M2 polarization. Interestingly, M2 polarization is not crucial to POI development as abrogation of monocyte infiltration did not prevent POI due to a compensation of the IL-10 loss by resident macrophages and neutrophils. Organ culture studies demonstrated that IL-10 deficiency impeded neutrophil migration toward the surgically traumatized ME. This mechanism is mediated by reduction of neutrophil attracting chemokines. Conclusion: Monocyte-derived macrophages are the major IL-10 source during POI. An IL-10 deficiency decreases the postoperative expression of neutrophil-recruiting chemokines, consequently reduces the neutrophil extravasation into the postsurgical bowel wall, and finally protects mice from POI.

Intestinal manipulation affects mucosal antimicrobial defense in a mouse model of postoperative ileus.

AIM: To explore the effects of abdominal surgery and interleukin-1 signaling on antimicrobial defense in a model of postoperative ileus. METHODS: C57BL/6 and Interleukin-1 receptor type I (IL-1R1) deficient mice underwent intestinal manipulation to induce POI. Expression of mucosal IL-1α, IL-1ß and IL-1R1 and several antimicrobial peptides and enzymes were measured by quantitative PCR or ELISA, western blotting or immunohistochemistry. Bacterial overgrowth was determined by fluorescent in-situ hybridization and counting of jejunal luminal bacteria. Translocation of aerobic and anaerobic bacteria into the intestinal wall, mesenteric lymph nodes, liver and spleen was determined by counting bacterial colonies on agar plates 48h after plating of tissue homogenates. Antimicrobial activity against E. coli and B. vulgatus was analyzed in total and cationic fractions of small bowel mucosal tissue homogenates by a flow cytometry-based bacterial depolarization assay. RESULTS: Jejunal bacterial overgrowth was detected 24h after surgery. At the same time point, but not in the early phase 3h after surgery, bacterial translocation into the liver and mesenteric lymph nodes was observed. Increased antimicrobial activity against E. coli was induced within early phase of POI. Basal antimicrobial peptide and enzyme gene expression was higher in the ileal compared to the jejunal mucosa. The expression of lysozyme 1, cryptdin 1, cryptdin 4 and mucin 2 were reduced 24h after surgery in the ileal mucosa and mucin 2 was also reduced in the jejunum. Postoperative IL-1α and IL-1ß were increased in the postoperative mucosa. Deficiency of IL-1R1 affected the expression of antimicrobial peptides during homeostasis and POI. CONCLUSION: Small bowel antimicrobial capacity is disturbed during POI which is accompanied by bacterial overgrowth and translocation. IL-1R1 is partially involved in the gene expression of mucosal antimicrobial peptides. Altered small bowel antimicrobial activity may contribute also to POI development and manifestation in patients undergoing abdominal surgery.

Advantages of laparoscopic compared to conventional surgery are not related to an innate immune response of peritoneal immune activation: an animal study in rats.

PURPOSE: Laparoscopic surgery (LS) has proved superior compared to conventional surgery (CS) regarding morbidity, length of hospital stay, rate of wound infection and time until recovery. An improved preservation of the postoperative immune function is assumed to contribute to these benefits though the role of the local peritoneal immune response is still poorly understood. Our study investigates the peritoneal immune response subsequent to abdominal surgery and compares it between laparoscopic and conventional surgery to find an immunological explanation for the clinically proven benefits of LS. METHODS: Wistar rats (N = 140) underwent laparoscopic cecum resection (LCR; N = 28), conventional cecum resection (CCR; N = 28), laparoscopic sham operation (LSO; N = 28), conventional sham operation (CSO; N = 28), or no surgical treatment (CTRL; N = 28). Postoperatively, peritoneal lavages were performed, leukocytes isolated and analyzed regarding immune function and phagocytosis activity. RESULTS: Immune function was inhibited postoperatively in animals undergoing LCR or CCR compared to CTRL reflected by a lower TNF-α (CTRL 3956.65 pg/ml, LCR 2018.48 pg/ml (p = 0.023), CCR 2793.78 pg/ml (n.s.)) and IL-6 secretion (CTRL 625.84 pg/ml, LCR 142.84 pg/ml (p = 0.009), CCR 169.53 pg/ml (p = 0.01)). Phagocytosis was not affected in rats undergoing any kind of surgery compared to CTRL. Neither cytokine secretion nor phagocytosis activity differed significantly between laparoscopic and conventional surgery. CONCLUSIONS: According to our findings the benefits associated with LS compared to CS cannot be explained by differences in the postoperative peritoneal innate immune response. Further studies are needed to elucidate the causes for a more favorable postoperative outcome in patients after LS compared to CS.

A role for 12/15-lipoxygenase-derived proresolving mediators in postoperative ileus: protectin DX-regulated neutrophil extravasation.

Resolution of inflammation is an active counter-regulatory mechanism involving polyunsaturated fatty acid-derived proresolving lipid mediators. Postoperative intestinal motility disturbances, clinically known as postoperative ileus, occur frequently after abdominal surgery and are mediated by a complex inflammation of the intestinal muscularis externa. Herein, we tested the hypothesis that proresolving lipid mediators are involved in the resolution of postoperative ileus. In a standardized experimental model of postoperative ileus, we detected strong expression of 12/15-lipoxygenase within the postoperative muscularis externa of C57BL/6 mice, predominately located within CX3CR1(+)/Ly6C(+) infiltrating monocytes rather than Ly6G(+) neutrophils. Mass spectrometry analyses demonstrated that a 12/15-lipoxygenase increase was accompanied by production of docosahexaenoic acid-derived lipid mediators, particularly protectin DX and resolvin D2, and their common precursor 17-hydroxy docosahexaenoic acid. Perioperative administration of protectin DX, but not resolvin D2 diminished blood-derived leukocyte infiltration into the surgically manipulated muscularis externa and improved the gastrointestinal motility. Flow cytometry analyses showed impaired Ly6G(+)/Ly6C(+) neutrophil extravasation after protectin DX treatment, whereas Ly6G(-)/Ly6C(+) monocyte numbers were not affected. 12/15-lipoxygenase-deficient mice, lacking endogenous protectin DX synthesis, demonstrated increased postoperative leukocyte levels. Preoperative intravenous administration of a docosahexaenoic acid-rich lipid emulsion reduced postoperative leukocyte infiltration in wild-type mice but failed in 12/15-lipoxygenase-deficient mice mice. Protectin DX application reduced leukocyte influx and rescued 12/15-lipoxygenase-deficient mice mice from postoperative ileus. In conclusion, our results show that 12/15-lipoxygenase mediates postoperative ileus resolution via production of proresolving docosahexaenoic acid-derived protectin DX. Perioperative, parenteral protectin DX or docosahexaenoic acid supplementation, as well as modulation of the 12/15-lipoxygenase pathway, may be instrumental in prevention of postoperative ileus.

Aggravation of Established Colitis in Specific Pathogen-free IL-10 Knockout Mice by Restraint Stress Is Not Mediated by Increased Colonic Permeability.

BACKGROUND: Pathogenic mechanisms responsible for the undulating symptom pattern, or indeed causative agents for the development, of inflammatory bowel diseases [IBD] are largely unknown. Many physicians and most patients are convinced that stress affects the course of IBD. As with most factors that contribute to IBD, it is unclear whether stress merely exacerbates established disease or indeed contributes to the development of disease. We designed this study to investigate whether stress induces or aggravates colitis in interkeukin-10 knockout [IL-10 ko] mice and to determine the role of intestinal permeability in this model of stress-related colitis. METHODS: The study was divided into two experiments depending on the age of the animals. Stress was induced by placing 5-week old disease-free mice or 8-week-old mice (IL-10ko and wild type [wt]) with mild colitis in movement restrainers for 2h twice daily for 7 days. The development of colitis was assessed clinically [weight and faecal pellet production], histologically [haematoxylin and eosin staining], and biochemically [colonic IL-2, IL-4, IL-6, IL-12p40, TNFα, and IFNγ]. Permeability was measured in Ussing chambers. RESULTS: Faecal pellet production increased significantly in all stressed animals compared with control animals, indicating successful application of stress. Stressed 8-week old mice lost weight [p < 0.001] and stressed IL-10(-/-) mice showed a significantly increased histological score compared with non-stressed or wt mice [p < 0.001]. There was no appreciable difference in cytokine production. Stress did not alter intestinal permeability. CONCLUSIONS: Restraint stress aggravates experimental colitis in 8-week old IL-10ko mice but cannot induce colitis in disease-free younger mice. This is not mediated by an increased intestinal permeability.

Postoperative ileus involves interleukin-1 receptor signaling in enteric glia.

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.

Surgical management of trigeminal neuralgia patients with recurrent or persistent pain despite three or more prior operations.

OBJECTIVE: To compare facial pain outcomes from different surgical techniques for patients with idiopathic trigeminal neuralgia (TN) who continue to have persistent or recurrent TN despite multiple operations. METHODS: Review of a prospective surgical database identified 62 TN patients with ongoing facial pain despite having undergone three or more prior operations (mean = 3.4) from July 1999 to March 2008. The mean patient age was 66.5 years and the mean pain duration was 11.8 years. Twenty-six patients (42%) underwent 33 additional procedures during the follow-up period. In total, 95 operations were performed: posterior fossa exploration (PFE; n = 37, 39%), stereotactic radiosurgery (n = 31, 33%), glycerol rhizotomy (n = 18, 19%), and balloon compression (n = 9, 10%). Follow-up (median = 35 months; range = 1 day to 103 months) was censored at time of subsequent surgery, last patient contact, or death. RESULTS: Complete pain relief after surgery (no pain, no medications) was 66% at 1 year and 50% at 3 years. Patients having PFE had better facial pain outcomes (70% complete relief at 3 years) compared to other procedures (36% at 3 years) (hazard ratio = 2.6, 95% CI = 1.3-5.1, P < .01). No difference was noted between radiosurgery and the percutaneous techniques. Additional surgery was performed in 8 patients after PFE (22%) compared to 25 patients (48%) after radiosurgery (n = 16), glycerol rhizotomy (n = 7), or balloon compression (n = 2; P = .02). CONCLUSIONS: Although no procedure is best for all patients, PFE gives the operating surgeon the option of performing either a nondestructive (microvascular decompression) or destructive (partial sensory rhizotomy) procedure and is associated with better facial pain outcomes for this difficult patient group.

Violence against women: the state of batterer prevention programs.

While both men and women can be victims, domestic violence usually consists of assaults on women, and most violence against women occurs within an intimate relationship. In the past twenty years, numerous state and provincial programs to intervene in domestic violence cases have developed. The programs tend to focus on treating batterers, although they also offer counseling to domestic violence victims. The jury remains out on the effectiveness of these programs. A major issue is whether the programs use appropriate standards. After an overview of the prevalence and nature of domestic violence, this article provides a discussion of those standards--their nature, effectiveness, and limitations. Another section discusses use of a batterer intervention program in an urban setting. Yet another section explores the implications of intimate partner violence and looks again at the effectiveness of batterer treatment within intervention programs. The article closes with a look at the way one state addresses domestic violence and treats it as a crime. An inescapable conclusion to be drawn from the discussion is that violence against women has its roots in cultural assumptions that must undergo change if the incidence of that violence is to be reduced.