The role of the vagus nerve: modulation of the inflammatory reaction in murine polymicrobial sepsis.

The particular importance of the vagus nerve for the pathophysiology of peritonitis becomes more and more apparent. In this work we provide evidence for the vagal modulation of inflammation in the murine model of colon ascendens stent peritonitis (CASP). Vagotomy significantly increases mortality in polymicrobial sepsis. This effect is not accounted for by the dilatation of gastric volume following vagotomy. As the stimulation of cholinergic receptors by nicotine has no therapeutic effect, the lack of nicotine is also not the reason for the reduced survival rate. In fact, increased septic mortality is a consequence of the absent modulating influence of the vagus nerve on the immune system: we detected significantly elevated serum corticosterone levels in vagotomised mice 24 h following CASP and a decreased ex vivo TNF-alpha secretion of Kupffer cells upon stimulation with LPS. In conclusion, the vagus nerve has a modulating influence in polymicrobial sepsis by attenuating the immune dysregulation.

Intraarterial recombinant tissue plasminogen activator thrombolysis of acute and semiacute lower limb arterial occlusion: quality assurance, complication management, and 12-month follow-up reinterventions.

OBJECTIVE: The purpose of this study is to assess the efficacy of intraarterial thrombolysis in acute and semiacute occlusions of the lower limb. MATERIALS AND METHODS: A total of 77 native arteries and 52 bypass grafts were investigated in 129 patients (mean [± SD] age, 64.6 ± 11.1 years) with acute (i.e., symptoms for ≤ 14 days) or semiacute (i.e., symptoms for > 14 days) peripheral arterial occlusions of the lower limb treated by catheter-directed recombinant tissue plasminogen activator (rt-PA) thrombolysis. Therapeutic success was retrospectively analyzed according to vessel type and duration of occlusion. Morbidity and mortality rates associated with thrombolytic treatment were calculated. The hospitalization period after primary intervention was recorded. Reinterventions and amputations were assessed at 12-month follow-up. RESULTS: Recanalization was accomplished by rt-PA thrombolysis in 73.6% of all cases. There was no difference in primary therapeutic success between native arteries and bypass grafts (p = 0.601). Thrombolysis was more effective in acute peripheral occlusions, and hospital stays were shorter than those for patients with semiacute occlusion (p = 0.001). The morbidity rate was 31% (minor complications, 20.2%; major complications, 10.9%), and the mortality rate was 2.3%. Within 12 months, radiologic and surgical interventions were necessary for 27 patients. The limb salvage rate after primarily successful recanalization was 89.5%. CONCLUSION: Intraarterial rt-PA thrombolysis is an effective and reasonable method for treating acute peripheral arterial occlusion. The method is less effective in semiacute occlusions, leading to extended hospitalization. Within 12 months, a quarter of the patients required reinterventions, and amputations were necessary in 10% of the cases.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) improves the innate immune response and enhances survival in murine polymicrobial sepsis.

OBJECTIVE: To investigate the role of tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) in postoperative polymicrobial abdominal sepsis.Sepsis is the leading cause of death among critically ill surgical patients. TRAIL is commonly known as an apoptosis-inducing agent in cancer cells. It also plays an important role in the regulation of inflammatory reactions. The role of TRAIL in polymicrobial sepsis is still unclear. DESIGN: Experimental animal model. SETTING: University laboratory. SUBJECTS: C57BL/6 mice. INTERVENTIONS: Colon ascendens stent peritonitis (CASP) was induced in female mice. One hour, 24 hrs, and 48 hrs after induction of CASP, murine recombinant TRAIL was given intravenously. MEASUREMENTS AND MAIN RESULTS: This study demonstrates a protective effect of TRAIL in CASP, an experimental model of murine polymicrobial sepsis. Intravenous administration of recombinant TRAIL to mice after CASP induction led to highly significantly prolonged survival. The migration of effector cells into the peritoneal cavity was strongly enhanced. Consequently, TRAIL-treated mice eliminated bacteria significantly better from the peritoneal cavity, the source of infection. Systemic spread of gut bacteria was also reduced by several orders of magnitude. As a result of the reduced systemic spread of bacteria, the accumulation of neutrophils within the spleen and mesenteric lymph nodes was strongly decreased. CONCLUSION: TRAIL-treated mice are highly protected from abdominal sepsis. Because diagnosis and therapy are frequently delayed in human sepsis, it is remarkable that TRAIL is effective when given via a therapeutic approach. Therefore, this study suggests a therapeutic potential for TRAIL in human sepsis. This should be addressed in future trials.

Implantation of alloplastic material increases survival of mice subsequently exposed to polymicrobial sepsis.

PURPOSE: Major surgery can modulate the immune system and by this the clinical course of following complications. Effects of minor surgical treatments on the immune system and septic complications are poorly understood. MATERIALS AND METHODS: We investigated the effect of a minor surgical procedure--the implantation of an osmotic pump--on the outcome of experimental polymicrobial sepsis (colon ascendens stent-induced peritonitis, CASP) in mice. RESULTS: Animals with pumps implanted 3 days prior to CASP showed an attenuated clinical course of sepsis and increased survival. While measured serum cytokine levels were not affected by the minor surgical stress of pump implantation, splenocyte secretion of IFN-gamma in response to lipopolysaccharide was increased. CONCLUSION: The early implantation of alloplastic material modulates the immune system and leads to an increased survival of a polymicrobial sepsis. Identifying the molecular nature of this effect might point the way to a new therapeutic approach to reduce sepsis mortality.

Selective depletion of alveolar macrophages in polymicrobial sepsis increases lung injury, bacterial load and mortality but does not affect cytokine release.

BACKGROUND: Resident tissue macrophages exert important functions during severe systemic infection and contribute to changes in local as well as systemic immune responses. Alveolar macrophages (AM) play a crucial role in airway diseases and in the defense against microorganisms invading the body via the bronchopulmonary tract. It has been postulated that AM are involved in the development of acute local disorders as a consequence of extrapulmonary stimuli like pancreatitis, peritonitis, or trauma. OBJECTIVE: The aim of this study was to analyze the local and systemic role of AM during sepsis using selective AM depletion in the murine colon ascendens stent peritonitis (CASP) model of polymicrobial sepsis. METHODS: 48 h prior to CASP surgery, AM of female C57BL/6 mice were selectively depleted by intratracheal application of clodronate liposomes (Lipo-clod). For control purposes, phosphate-buffered saline (PBS) liposomes (Lipo-PBS) were used. RESULTS: CASP led to significantly elevated levels of local and systemic cytokines independent of the presence of AM. In contrast, levels of gut-derived bacteria in bronchoalveolar lavage and lung of septic mice were significantly higher in Lipo-clod-treated animals compared to Lipo-PBS-treated animals. After CASP-induced sepsis, local barrier dysfunction in the lung was detected; AM depletion resulted in severely enhanced development of acute lung injury. Consequently, Lipo-clod-treated animals showed strongly reduced survival rates after CASP. CONCLUSIONS: Contrarily to other macrophage populations, AM do not significantly contribute to local and systemic cytokine release during polymicrobial abdominal sepsis. AM have important protective functions for local clearance of gut-derived bacteria and attenuation of lung injury.

Detrimental role of CC chemokine receptor 4 in murine polymicrobial sepsis.

CC chemokine receptor 4 (CCR4) and its two ligands, CCL17 and CCL22, are critically involved in different immune processes. In models of lipopolysaccharide-induced shock, CCR4-deficient (CCR4(-/-)) mice showed improved survival rates associated with attenuated proinflammatory cytokine release. Using CCR4(-/-) mice with a C57BL/6 background, this study describes for the first time the role of CCR4 in a murine model of polymicrobial abdominal sepsis, the colon ascendens stent peritonitis (CASP). CASP-induced sepsis led to a massive downregulation of CCR4 in lymphoid and nonlymphoid tissues, whereas the expression of CCL17 and CCL22 was independent of the presence of CCR4. After CASP, CCR4(-/-) animals showed a strongly enhanced bacterial clearance in several organs but not in the peritoneal lavage fluid and the blood. In addition, significantly reduced levels of proinflammatory cytokines/chemokines were measured in organ supernatants as well as in the sera of CCR4(-/-) mice. CCR4 deficiency consequently resulted in an attenuated severity of systemic sepsis and a strongly improved survival rate after CASP or CASP with intervention. Thus, our data provide clear evidence that CCR4 plays a strictly detrimental role in the course of polymicrobial sepsis.

Sepsis affects cardiac expression of multidrug resistance protein 5 (MRP5, ABCC5), an ABC-type CGMP export pump.

One of the clinical characteristics associated with septic shock is heart failure. Several lines of evidence indicate that functional consequences of heart failure in septic shock are linked to the activated NO-cyclic guanosine monophosphate (NO-cGMP) pathway. We have previously shown that the high-affinity cGMP export transporter, multidrug resistance protein 5 (MRP5), is expressed in the heart, which modulates intracellular concentrations and, hence, the effects of cGMP. Thus, modified expression of cardiac MRP5 in septic shock can alter cGMP concentrations and contribute to the development of heart failure. We therefore investigated MRP5 expression in the heart using two established murine models of septic shock (intraperitoneal LPS injection and surgical implantation of a stent into the ascending colon, resulting in a multibacterial peritonitis [CASP, colon ascendens stent peritonitis] in C57BL/6N mice, respectively; n = 38). Cardiac MRP5 was assessed by quantitative polymerase chain reaction and immunofluorescence. The protein was localized in the endothelial wall, smooth muscle, and cardiac myocytes. MRP5 mRNA expression was significantly reduced compared with controls both in the LPS (31.9 +/- 16.8 x 10(-4) vs. 54.1 +/- 14.8 x 10(-4), P = 0.025) and CASP model (18.3 +/- 9.4 x 10(-4) vs. 42.8 +/- 12.1 x 10(-4), P = 0.009; MRP5/glyceraldehyde 3-phosphate dehydrogenase copy numbers, respectively). In parallel, IL-6 plasma levels were significantly increased in both models. Incubation of cultured murine cardiomyocytes (HL1) with 5 ng/mL IL-6 resulted in decreased expression of MRP5 (54% of control), as did incubation of the cells with serum from septic mice (LPS serum, 22% of control; CASP serum, 11% of control). In conclusion, cardiac expression of the cGMP export transporter MRP5 is decreased in two murine models of septic shock, most likely by a transcriptional mechanism. Reduced cGMP export as a consequence of decreased MRP5 expression can attenuate heart failure in sepsis.

Cecal ligation and puncture versus colon ascendens stent peritonitis: two distinct animal models for polymicrobial sepsis.

Colon ascendens stent peritonitis (CASP) and cecal ligation and puncture (CLP), two animal models designed to closely mimic the clinical course of intra-abdominal sepsis, were compared. In the past, immunomodulatory therapies developed in animal studies failed to be successful in humans. As a consequence, the established animal sepsis models were criticized. It has been proposed that present models had to be reevaluated, and new, clinically more relevant models should be evolved. CLP procedure was performed puncturing once (CLP[1]) or twice (CLP[2]) the ligated cecum of C57BL/6 mice. In the CASP model, a stent with defined diameter was surgically inserted into the ascending colon. Survival, bacterial load, immunohistochemistry, and serum cytokine levels were analyzed in the groups. Survival after CASP procedure correlated strongly with the stent diameter, whereas the number of punctures in CLP did not significantly change survival rate. Bacterial loads of peritoneal lavage, liver, and lung, as well as serum cytokine levels (tumor necrosis factor, interleukin 1 beta, interleukin 10) steadily increased from 6 to 24 h after the CASP procedure. In contrast, continuously low amounts of bacteria and cytokines were found in CLP mice at any point of time. Twenty-four hours after CLP surgery, the ligated cecum was covered by adhesive small bowel loops, whereas in CASP mice, the intestinal leakage was then still present. The CASP model mimics closely the clinical course of diffuse peritonitis with early and steadily increasing systemic infection and inflammation (systemic inflammatory response syndrome). In contrast, CLP reveals a model of intra-abdominal abscess formation with sustained and minor signs of systemic inflammation.

Rare case of a large mediastinal cyst involved in Hodgkin's lymphoma.

We report a case of a mediastinal cystic retrosternal process, discovered by magnetic resonance imaging (MRI) in a 19-year-old male patient, with unusual inhomogenous signals in both T1- and T2-weighted images and contrast-enhancing septation. Macroscopically, the tumor weighed 1330 g, and was constituted by one dominating cyst measuring 14 cm in diameter. Additional small cysts were seen microscopically. The cystic wall was continuously infiltrated by nodular sclerosing Hodgkin's lymphoma, also affecting adjacent lymph-nodes. Age and sex of the patient and the diagnosed subtype of Hodgkin's lymphoma are in line with previously reported rare cases of mediastinal cysts with Hodgkin's lymphoma. The cyst reported here, most likely a secondary thymic cyst, is larger than those reported before. The main reason for the development of these cysts might be the accompanying inflammation of the lymphoma. Little is known about the imaging features of mediastinal cysts caused by lymphoma. Plain thymic cysts are normally homogenous on T1- and T2-weighted images. Hodgkin's lymphoma might be homogenous on T1-weighted images and is mostly inhomogenous on T2-weighted images. In case of inhomogenous cysts with contrast-enhancing septation, one should consider the diagnosis of an associated neoplasm.

Colon ascendens stent peritonitis (CASP)--a standardized model for polymicrobial abdominal sepsis.

Sepsis remains a persistent problem on intensive care units all over the world. Understanding the complex mechanisms of sepsis is the precondition for establishing new therapeutic approaches in this field. Therefore, animal models are required that are able to closely mimic the human disease and also sufficiently deal with scientific questions. The Colon Ascendens Stent Peritonitis (CASP) is a highly standardized model for polymicrobial abdominal sepsis in rodents. In this model, a small stent is surgically inserted into the ascending colon of mice or rats leading to a continuous leakage of intestinal bacteria into the peritoneal cavity. The procedure results in peritonitis, systemic bacteraemia, organ infection by gut bacteria, and systemic but also local release of several pro- and anti-inflammatory cytokines. The lethality of CASP can be controlled by the diameter of the inserted stent. A variant of this model, the so-called CASP with intervention (CASPI), raises opportunity to remove the septic focus by a second operation according to common procedures in clinical practice. CASP is an easily learnable and highly reproducible model that closely mimics the clinical course of abdominal sepsis. It leads way to study on questions in several scientific fields e.g. immunology, infectiology, or surgery.

Kupffer cell depletion reduces hepatic inflammation and apoptosis but decreases survival in abdominal sepsis.

OBJECTIVE: During abdominal sepsis, the activation of hepatic Kupffer cells (KC) and its consequences are of central interest. This study evaluates the impact of selective KC depletion on hepatic microcirculation, cytokine release, and systemic alterations in the colon ascendens stent peritonitis (CASP), a model of polymicrobial abdominal sepsis. METHODS: For KC depletion clodronate liposomes were injected 24 h before CASP surgery in female C57BL/6N mice. Three and 12 h after CASP, in-vivo fluorescence microscopy of the liver was performed. Analysis of hepatocellular apoptosis was conducted by immunohistochemistry. In addition, levels of tumor necrosis factor (TNF), IL-6, and IL-10 in the liver, lungs, spleen, and plasma were determined, and bacteriology and survival analysis were performed. RESULTS: CASP led to significant sinusoidal perfusion failure, increased leukocyte recruitment, hepatocellular apoptosis and increased levels of TNF, IL-6, and IL-10 in the liver and plasma. KC depletion before CASP significantly reduced leukocyte recruitment to the liver and hepatocellular apoptosis. IL-10 secretion decreased dramatically in the liver and plasma of KC-depleted septic mice. In contrast, TNF levels were clearly elevated after clodronate treatment. In the lung and spleen, a compensatory upregulation of IL-10 could be detected after KC depletion. Clodronate treatment resulted in a significant reduction in survival. CONCLUSION: The results indicate that KC depletion is locally protective in polymicrobial abdominal sepsis, as it reduces hepatic inflammation and apoptosis. These effects could be observed in the presence of clearly elevated TNF levels. However, the lack of IL-10 in KC-depleted mice resulted in a detrimental systemic proinflammation.

Seasonal variations in inflammatory responses to sepsis and stress in mice.

OBJECTIVE: In this study, we analyzed seasonal variations of immunoreactivity using a model of septic shock and a model of immunosuppression induced by chronic stress in mice. DESIGN: Retrospective comparative study using animals of experiments performed between 2001 and 2006 to identify seasonal variations in inflammatory responsiveness of mice. SETTING: University-based research laboratory. SUBJECTS: C57Bl/6 mice and BALB/c mice. INTERVENTIONS: For analyzing septic shock, we used the hyperinflammatory model of colon ascendens stent peritonitis. Immunosuppression was induced by 4.5 days of intermittent combined acoustic and restraint stress. MEASUREMENTS AND MAIN RESULTS: We show that mice kept with 12:12-hr light/dark rhythm had an enhanced risk to die of experimentally-induced hyperinflammatory peritonitis performed in summer or autumn compared with the other seasons. This finding was associated with an exaggerated proinflammatory response of C57Bl/6 mice in summer or autumn compared with moderate inflammatory reactivity in winter and spring. Consistent with these results, we report that the severity of a stress-induced immunosuppression is less pronounced in BALB/c mice that were exposed to chronic psychological stress in the summer compared with exposure in winter. Coping with chronic psychological stress of these animals was correlated with less pronounced corticosterone release, less severe lymphocytopenia, and a lower ex vivo inducibility of interleukin-10, thereby attenuating a stress-mediated immunosuppressive state. Mice subjected to chronic stress in the summer season showed increased coping compared with mice that were stressed in the winter season. CONCLUSIONS: Our results suggest that seasonal changes of the host's hypothalamus-pituitary-adrenal axis response influence the risk of infection and the susceptibility to stress, which interferes with the outcome after infection.

Impact of interleukin-12, oxidative burst, and iNOS on the survival of murine fecal peritonitis.

Abdominal sepsis due to secondary fecal peritonitis following anastomosis insufficiency is a rare but life threatening complication of colorectal surgery. The induction of IFN-gamma by IL-12 is believed to play a key role in sepsis as it promotes antibacterial effector mechanisms such as oxidative burst or nitric oxide induction. The impact of gene deficiency for IL-12 (IL-12p40 KO), oxidative burst (p47(phox) KO), or NO induction (iNOS KO) on the outcome of fecal peritonitis was characterized using the murine Colon Ascendens Stent Peritonitis model (CASP). In the IL-12p40 KO model, 3 and 12 h after surgery, serum cytokine levels of IL-1beta, TNF, IL-18, and IL-10 were analyzed. Expression of IL-1beta, IL-10, IP-10, and MIP-1alpha was measured in lung and liver by RNAse Protection Assay. IL-12p40 and iNOS-deficient mice exhibited a significantly higher susceptibility to CASP as compared to the controls, whereas no significant difference was observed in p47(phox) KO mice. Absence of IL-12 resulted in delayed expression of proinflammatory cytokines and chemokines in both the liver and the lung, and was associated with significant reduction of IL-1beta levels in the serum 12 h after CASP. IL-12 and iNOS possess protective functions in fecal murine peritonitis. Surprisingly, no significant contribution of oxidative burst to the immune response was observed. Overall, these findings suggest that IL-12 deficiency causes a profound delay of the immune response after polymicrobial challenge resulting in significantly increased susceptibility in the CASP model.

The vagal nerve as a link between the nervous and immune system in the instance of polymicrobial sepsis.

BACKGROUND: The role of the vagal nerve in the autonomic nervous system is widely well known. Recently, an additional function was revealed serving as a connector between the nervous and immune system. This connection is called the "cholinergic inflammatory pathway." Through stimulation of the acetylcholine receptors located upon the macrophages, the "unspecific" immune system can be directly influenced. METHODS: The vagal nerve was completely transected directly posterior to its passage through the diaphragm. The effect of complete vagotomy was analyzed using a murine model of polymicrobial peritonitis (colon ascendens stent peritonitis, CASP). Survival and clinical course of vagotomized or sham-operated mice were analyzed in the CASP model. RESULTS: After CASP surgery, vagotomy led to a significantly increased mortality (64.7%) in comparison to sham-vagotomized animals (34%). No difference in the bacterial load of various tissues (lung, liver, spleen, blood, lavage fluid, and kidney) from septic animals with or without vagotomy was observed. Vagotomized animals reveal elevated serum cytokine levels (TNF, IL-6, IL-10, and MCP-1) 20 h after the induction of polymicrobial peritonitis. CONCLUSION: The vagal nerve is therefore an important modulator of the immune system.

CCR4-deficient mice show prolonged graft survival in a chronic cardiac transplant rejection model.

Chronic graft rejection mediated by cellular immune responses still poses a serious clinical problem in transplant surgery. Chemokines coordinate the recruitment of leukocytes in inflammatory and immune responses. Their precise functions in the rejection of allografts are still ill defined. This study investigates the role of chemokine receptor 4 (CCR4) in acute and chronic cardiac allograft rejection in mice. Allogeneic hearts were transplanted into CCR4 deficient (CCR4(-/-)) and control recipients. Reverse transcription-PCR showed transcription of macrophage-derived chemokine and thymus and activation-regulated chemokine, the cognate chemokine ligands of CCR4, within the graft. Compared to wild-type controls, acute allograft rejection in CCR4(-/-) recipients was only slightly prolonged. In contrast, in a gallium nitrate chronic cardiac allograft rejection model, cardiac graft survival was significantly prolonged in CCR4(-/-) recipients. A relative increase in the percentage of graft infiltrating CD8(+) T cells in CCR4(-/-) recipients was observed 30 days after transplantation and was accompanied by a decrease in CD4(+) T cells. Moreover, the percentage of NK1.1(+)CD3(+) graft-infiltrating cells was significantly reduced on day 5 and day 30 post transplantation. These findings indicate that CCR4 is involved in the recruitment of NK1.1(+)CD3(+) cells into cardiac allografts and clearly establish an important and novel role for CCR4 in chronic graft rejection.