Clostridium difficile toxins influence hepatocyte protein synthesis through the interleukin 1 receptor.

HYPOTHESIS: Clostridium difficile toxins require interleukin 1 (IL-1) production or a functioning IL-1 receptor to elicit acute-phase protein production by murine hepatocytes. DESIGN: Experimental study. SETTING: Research laboratory at the DVA Medical Center, St Louis, Mo. CELLS STUDIED: Hepatocytes prepared from normal mice, from knockout mice deficient in IL-1 production due to loss of IL-1 converting enzyme, or from knockout mice deficient in the IL-1 p80 receptor. INTERVENTIONS: Cells were treated with lipopolysaccharide, a crude C difficile toxin extract, or purified C difficile toxins A or B for 24 hours in vitro, then radiolabeled with (35)S methionine. Newly synthesized acute-phase proteins were identified by electrophoresis and autoradiography. MAIN OUTCOME MEASURES: Synthesis of a 23-kd acute-phase protein in response to the various stimuli. RESULTS: Lipopolysaccharide, C difficile culture extract, and purified toxins A and B stimulated the synthesis of the 23-kd acute-phase protein by hepatocytes from normal mice and by hepatocytes from knockout mice deficient in the IL-1 converting enzyme. However, hepatocytes from knockout mice deficient in the IL-1 p80 receptor failed to produce this acute-phase protein when treated with the C difficile toxins, although they responded fully to lipopolysaccharide. CONCLUSIONS: Stimulation of acute-phase protein synthesis by C difficile toxins does not require IL-1 production, but does require a functioning IL-1 p80 receptor. This suggests that some of the actions of these toxins are mediated by this receptor.

Increased small bowel epithelial turnover in interleukin-1 receptor knockout mice.

OBJECTIVE: To determine whether interleukin-1 (IL-1) affects the cellular homeostasis of small bowel mucosa, the authors studied apoptosis and proliferation in small bowel epithelium in two groups of C57 mice: an IL-1 receptor knockout group, and a control wild-type group. SUMMARY BACKGROUND DATA: Gut mucosal integrity is maintained by a balance of cell proliferation and cell death. Recent reports suggest that IL-1, a proinflammatory cytokine, increases cell death by apoptosis in some epithelial cells. METHODS: Twenty-four male C57BL6 IL-1 receptor (type I) knockout mice were killed, and small bowel was removed for study. Twenty-four wild-type mice (C57-BL6) served as controls. Body weights, bowel length, and mucosal morphology were examined for phenotypic differences. Apoptosis was quantified by terminal deoxyuridine nick-end labeling (TUNEL) immunohistochemical staining and cellular proliferation by proliferation cell nuclear antigen staining. Whole mucosal protein was analyzed for nuclear factor-kappaB expression. Groups were analyzed by t test. RESULTS: The absence of IL-1 type I receptor in knockout mice was verified by reverse transcriptase-polymerase chain reaction. IL-1 receptor null mice were larger than wild-type controls, with a longer small bowel; however, the index of small bowel length to total body weight did not differ between groups. The percentage of apoptotic cells was higher in IL-1 receptor null mice than in wild-type mice; the proliferation index also increased. Mucosal height and other measures of mucosal morphology were not different. Genotypic absence of IL-1 receptors was associated with decreased expression of nuclear factor-kappaB in whole mucosal protein extracts. CONCLUSIONS: Both apoptosis and proliferation increased in gut epithelial cells of mice without IL-1 receptors, suggesting increased cell turnover with no change in net balance. This model represents an opportunity to examine potential mechanisms of gut epithelial turnover in vivo, under both normal conditions and in conditions of mucosal proliferation and atrophy.

Involvement of p38 mitogen-activated protein kinase in the induction of tolerance to hemorrhagic and endotoxic shock.

BACKGROUND: Exposure to sublethal hemorrhage (SLH) makes rats tolerant to subsequent hemorrhagic or septic shock and is associated with altered NF-kappaB activity. The purpose of this study was to explore whether changes in p38 mitogen-activated protein (MAP) kinase activity also occur in the induction of tolerance by SLH. METHODS: Rats were made tolerant by SLH or sham operation. Twenty-four hours later rats were exposed to lipopolysaccharide (LPS) or had peritoneal macrophages (Mphi) isolated. CNI-1493, a p38 MAP kinase inhibitor, or saline was given prior to SLH. Lungs were harvested 1 h after SLH or LPS and total protein was extracted. Peritoneal Mphi were stimulated with LPS (10 microg/ml) and total protein was isolated 1 h later. Active, dually phosphorylated p38 MAP kinase was determined by Western blot. Tumor necrosis factor (TNF) was measured in Mphi supernatants by enzyme-linked immunosorbent assay (ELISA) 18 h after LPS. RESULTS: SLH activated p38 MAP kinase in the lung and this was inhibited by CNI-1493. Twenty-four hours later, lung p38 MAP kinase activity increased to the same degree in tolerant and sham rats following LPS, but much more prominently in the CNI-1493 treated rats. There was no p38 activity in peritoneal Mphi at baseline, and similar to lung p38, LPS led to increased p38 activity which was most significant in Mphi from rats that received CNI-1493 prior to SLH. TNF production by tolerant Mphi in response to LPS was significantly (P < 0.05, t test) decreased and p38 inhibition with CNI-1493 at the time of SLH reversed the inhibitory effects of tolerance on TNF production. CONCLUSIONS: TNF production by tolerant Mphi following a second insult (LPS) is attenuated despite preservation of normal p38 MAP kinase activity. However, activation of this intracellular second messenger is a necessary step in the "cellular reprogramming" that occurs during the induction of tolerance by SLH.

Clostridium difficile toxin: cytoskeletal changes and lactate dehydrogenase release in hepatocytes.

BACKGROUND: We have found that Clostridium difficile toxins can evoke hepatocyte acute-phase protein synthesis, and that this effect is dependent on a functioning interleukin-1 (IL-1) receptor. The present study was undertaken to determine if C. difficile toxicity, as determined by actin rearrangement and lactate dehydrogenase (LDH) release, also requires a functioning IL-1 receptor. METHODS: Primary hepatocyte cultures were prepared from normal mice, knockout mice deficient in the IL-1-converting enzyme (ICE), and knockout mice deficient in the IL-1 p80 receptor. Hepatocytes were treated for 24 h with C. difficile culture extract, purified C. difficile toxin A, or purified C. difficile toxin B. The actin cytoskeleton was examined using confocal microscopy, and LDH release was measured by spectrophotometric analysis. RESULTS: C. difficile culture extract, toxin A, and toxin B induced collapse of the actin cytoskeleton in hepatocytes from normal mice. Hepatocytes from both the ICE-deficient mice and the IL-1 p80 receptor-deficient mice demonstrated similar responses to both toxins. These toxins also induced significant LDH release in a concentration-dependent fashion in the normal hepatocytes and the ICE-deficient hepatocytes. However, no significant increase in LDH release was observed in hepatocytes from IL-1 p80 receptor-deficient mice. CONCLUSIONS: C. difficile toxins induce actin cytoskeletal collapse independent of IL-1 or the IL-1 receptor. In contrast, toxin-stimulated LDH release was dependent on the presence of the IL-1 receptor. Thus, separate pathways appear to mediate toxic effects as manifested by actin rearrangement and LDH release.

The false-positive parathyroid sestamibi: a real or perceived problem and a case for radioguided parathyroidectomy.

OBJECTIVE: To demonstrate that the positive parathyroid sestamibi scan, if correctly interpreted and applied, truly represents a parathyroid adenoma, never a "false-positive" scan. SUMMARY BACKGROUND DATA: Although the sestamibi scan is widely ordered preoperatively to locate parathyroid adenomas, concern about a false-positive scan often causes surgeons to distrust the results. Tissues such as thyroid adenomas and lymph nodes have been blamed for false-positive studies, but the radioactivity of these presumed false-positive tissues has never been measured. METHODS: Over an 1 8-month period, 17 patients were referred for persistent primary hyperparathyroidism after undergoing at least one neck exploration. All patients had a sestamibi scan prior to their initial operation that was interpreted as clearly positive and then, during or after an unsuccessful operation, deemed false-positive by the surgeon. At the authors' institution, all patients underwent repeat sestamibi scintigraphy and were taken to the operating room while radioactive for a minimally invasive radioguided parathyroidectomy (MIRP). RESULTS: The authors' sestamibi scans demonstrated the same single focus of radioactivity displayed on the outside scans, clearly positive. During MIRP, an adenoma was successfully located and removed in all patients, with confirmation of the diagnosis by quantitative differential radioactivity and subsequent histologic examination. Removal of the radioactive tissue cured all patients. CONCLUSION: Intraoperative nuclear mapping permitted identification and removal of parathyroid adenomas in all patients with positive sestamibi scans that had previously been labelled false-positive, indicating that each patient would have been cured during their previous operation if radioguided techniques were used. Surgeons should be extremely cautious in deciding intraoperatively that a positive sestamibi scan is a false-positive scan.

Tolerance to shock: an exploration of mechanism.

OBJECTIVE: To determine if cross-tolerance to septic shock could be induced by a previous insult with sublethal hemorrhage (SLH) and to characterize the mechanisms involved in this induced protective response. BACKGROUND DATA: It is possible to condition animals by prior SLH such that they tolerate an otherwise lethal hemorrhage. It is also possible to condition animals with low doses of lipopolysaccharide (LPS) so that they survive a "lethal" septic insult. However, a paucity of information exists on cross-tolerance between hemorrhage and sepsis. METHODS: Rats were made tolerant by conditioning SLH or sham operation. Twenty-four hours later, tolerant and sham rats were exposed to a lethal dose of LPS. To explore the mechanism of tolerance induction, rats were given the macrophage (Mphi) inhibitor CNI-1493 or saline carrier before SLH. Survival and pulmonary vascular injury were determined after LPS. Serum tumor necrosis factor (TNF) levels and splenic Mphi TNF gene expression were measured at several time points. RESULTS: Prior SLH indeed made rats tolerant and imparted a significant survival benefit and reduction in pulmonary vascular injury after LPS. The tolerance induced by SLH was reversed by Mphi inhibition. Tolerant animals had low serum TNF levels immediately after SLH and reduced circulating TNF levels after LPS. SLH, however, did not inhibit the augmentation of TNF gene expression after LPS. CONCLUSIONS: Sublethal hemorrhage bestows protection against a lethal LPS challenge. Inhibition of the Mphi attenuated the benefit of the tolerance induced by SLH. Circulating TNF but not TNF gene after LPS is lessened by SLH. This implicates changes in Mphi intracellular signaling in induction of the tolerant state.

Sublethal hemorrhage blunts the inflammatory cytokine response to endotoxin in a rat model.

BACKGROUND: Tolerance to lipopolysaccharide (LPS) induced by previous hemorrhage in mice is associated with a blunted interleukin 1 (IL-1) response, suggesting down-regulation of the cytokine cascade as a possible protective mechanism. This study was undertaken to determine whether prehemorrhage induces attenuation of the cytokine response to sepsis beyond IL-1 in a rat model and whether this response occurs at the level of gene transcription. METHODS: Sprague-Dawley rats underwent sublethal hemorrhage, lethal intraperitoneal endotoxin, or sublethal hemorrhage with delayed lethal endotoxin. Animals were killed 12 hours after LPS injection or 24 hours after hemorrhage. IL-1 and tumor necrosis factor (TNF) mRNA levels were determined on total splenic RNA using reverse-transcriptase polymerase chain reaction, and serum cytokine levels were determined using enzyme-linked immunosorbent assay. RESULTS: Animals that received LPS alone mounted an IL-1 and TNF response (RNA and protein) much higher than animals subjected to hemorrhage alone. TNF and IL-1 gene expression and protein levels in prehemorrhaged animals that received LPS, however, were significantly lower than those of animals that received LPS alone. CONCLUSION: Hemorrhage induces early IL-1 and TNF gene expression, which blunts their subsequent expected increase after endotoxic challenge. These findings validate previously documented immune-modulated protective effects of the first insult in a two-hit model.

New approaches to acute pancreatitis: role of inflammatory mediators.

Regardless of whether the initiating event is alcohol, gallstones or some other less common cause, acute pancreatitis progresses in a predictable manner eventually leading to failure of multiple unrelated organs. Very quickly following the inciting event, a local inflammatory process is initiated which results in the local production of inflammatory mediators. Virtually all patients with acute pancreatitis will experience some symptoms related to this local inflammation, with some resolving completely at this point. Most patients will go on to develop a systemic hyperinflammatory state expressed as the development of fever, tachycardia, tachypnea, and mild acid-base disturbances. Although this systemic hyperinflammatory state is usually mild, occasionally it may be very severe resulting in overt distant organ failure. With a more thorough understanding of the inflammatory mediators responsible for this hyperinflammatory state, many new therapeutic approaches are on the horizon.

Hemolytically active (acylated) alpha-hemolysin elicits interleukin-1beta (IL-1beta) but augments the lethality of Escherichia coli by an IL-1- and tumor necrosis factor-independent mechanism.

Many pathogenic Escherichia coli produce the toxin alpha-hemolysin (Hly), and lipopolysaccharide (LPS), interleukin-1 (IL-1), and tumor necrosis factor (TNF) have all been recognized as important effector molecules during infections by gram-negative organisms. Despite the characterization of many in vitro effects of hemolysin, no direct relationship has been established between hemolysin, LPS, proinflammatory cytokine production, and E. coli-induced mortality. Previously, we have shown in vivo that hemolysin elicits a distinct IL-1alpha spike by 4 h into a lethal hemolytic E. coli infection. Using three transformed E. coli strains, WAF108, WAF270, and WAH540 (which produce no Hly [Hlynull], acylated Hly [Hlyactive], or nonacylated Hly [Hlyinactive], respectively), we sought to determine the specific roles of hemolysin acylation, LPS, IL-1, and TNF in mediating the lethality of E. coli infection in mice. WAF270 was 100% lethal in BALB/c, C3H/HeJ, and C57BL/6 mice; in mice pretreated with antibody to the type 1 IL-1 receptor; in type 1 IL-1 receptor-deficient mice; and in dual (type 1 IL-1 receptor-type 1 TNF receptor)-deficient mice at doses which were nonlethal (0%) with both WAF108 and WAH540. At lethal doses, WAF270 killed by 6 +/- 2.3 h while WAF108 and WAH540 killed at 36 +/- 9.4 and 36 +/- 13.8 h, respectively. These differences in mortality were not due to IL-1 or TNF release, and the enhanced expression of LPS, which corresponded to Hly expression, was not likely the primary factor causing mortality. We demonstrate that bacterial fatty acid acylation of hemolysin is required in order for it to elicit IL-1 release by monocytes and to confer its virulence on E. coli.

Cytokine activation through sublethal hemorrhage is protective against early lethal endotoxic challenge.

OBJECTIVES: To determine the immunologic consequences of nonlethal hemorrhage on subsequent exposure to lipopolysaccharide (LPS) and to determine the role of interleukin 1 beta (IL-1) specifically in mediating the response to LPS with and without prior hemorrhage. DESIGN: Prospective, randomized, controlled experimental trial. PARTICIPANTS: Male BALB/c mice and transgenic mice deficient in IL-1 converting enzyme. INTERVENTIONS: Animals were subjected to hemorrhage (by cardiac puncture), LPS challenge by intraperitoneal injection, or hemorrhage followed 24 hours later by LPS challenge. Mortality was assessed every 4 hours for 96 hours following hemorrhage or LPS exposure. Serum IL-1 levels were determined 24 hours after exposure to hemorrhage and LPS. SETTING: University of South Florida Core General Surgery Research Facility, Tampa. MAIN OUTCOME MEASURES: Mortality and serum IL-1 levels. RESULTS: Hemorrhage alone resulted in complete survival, whereas LPS alone resulted in near-complete (95%) mortality. Hemorrhage, when given 24 hours before LPS challenge, afforded significant protection compared with LPS alone (67% survival vs 5% survival; P < .001). Serum IL-1 levels 24 hours after exposure to LPS were significantly lower in prehemorrhaged mice than in those receiving LPS alone. Transgenic mice incapable of producing biologically active IL-1 were further protected, demonstrating near-complete (95%) survival following hemorrhage and LPS challenge. CONCLUSIONS: Cytokine activation through nonlethal hemorrhage attenuates subsequent IL-1 response to early immunologic challenge. Such immune suppression appears to be protective early on and is supported by the near-complete immunity to LPS in animals incapable of producing biologically active IL-1.

Tissue-specific cytokine production during experimental acute pancreatitis. A probable mechanism for distant organ dysfunction.

Our purpose was to determine if cytokines are produced systemically during acute pancreatitis. Proinflammatory cytokines are elevated during acute pancreatitis and have been implicated in the progression of pancreatitis-associated multiple organ dysfunction. Whether these mediators are produced within all tissues or very few specific organs is not known. Edematous pancreatitis was induced in adult male mice by IP injection of cerulein. Necrotizing pancreatitis was induced in young female mice by feeding a choline-deficient, ethionine supplemented diet. Animals were sacrificed as pancreatitis worsened, with multiple organs prepared for tissue mRNA and protein analysis by RT-PCR and immunoblotting. Pancreatitis severity was established by histologic grading and serum amylase and lipase. There was no cytokine mRNA or protein detectable prior to the induction of pancreatitis. Tumor necrosis factor-alpha (TNF-alpha) and interleukin-1-beta (IL-1 beta) mRNA and protein were detected within the pancreas early in the course of pancreatitis in both models, coinciding with the development of hyperamylasemia (both P < 0.001). Interleukin-6 was produced in the pancreas after pancreatitis was more fully developed (P < 0.001). IL-1 beta and TNF-alpha were subsequently produced in large amounts in lung, liver, and spleen but never within kidney, cardiac muscle, or skeletal muscle. A significant delay between pancreatic and distant organ cytokine production was always observed. It is concluded that proinflammatory cytokines are produced within the pancreas and within organs known to develop dysfunction during severe pancreatitis. Cytokine production is tissue specific, correlates with disease severity, and occurs within the pancreas first and subsequently within distant organs.

Matrix metalloproteinase inhibition attenuates human pancreatic cancer growth in vitro and decreases mortality and tumorigenesis in vivo.

Matrix metalloproteinases (MMP) are enzymes responsible for extracellular matrix degradation, a critical component influencing the growth and metastatic potential of cancer. The purpose of this study was to determine the in vitro effects of MMP inhibition on human pancreatic cancer cells and to document its effect on cancer growth in vivo. The effect of MMP inhibition was determined using the MMP inhibitor BB-94 and a moderately differentiated pancreatic cancer cell line (HPAC). In vitro, a dose response curve was generated over 5 days utilizing the MTT [3(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay. In vivo, using an established orthotopic model for pancreatic cancer (LD100 = 80 days), 22 nude mice with orthotopic tumors (30 were implanted) received either BB-94 or vehicle beginning 4 days prior to implantation and continuing to death or sacrifice on Day 70. Mice were weighted weekly. At death/sacrifice, tumors were weighted, volume determined, and metastases/ distant spread documented. In vitro, BB-94 had little effect on HPAC proliferation at 40 ng/ml but achieved progressively greater to near complete inhibition at doses up to 4000 ng/ml while maintaining cell viability. In vivo, BB-94 significantly increased length of survival (69 +/- 0.1 days vs. 56 +/- 3.1 days) and necropsy weight (25.7 +/- 1.67 g vs. 19.8 +/- 1.14 g) while decreasing metastatic rate (1 vs. 20) and tumor size (0.14 +/- 0.02 g vs. 0.65 +/- 0.1 g). MMP inhibition limits HPAC proliferation in a dose-dependent fashion without direct cytotoxic effects in vitro. Mice harboring orthotopic tumors treated with BB-94 demonstrated significant reductions in tumor weight, volume, and metastases which corresponded to increased animal weight and prolonged survival.

Endothelial structural integrity is maintained during endotoxic shock in an interleukin-1 type 1 receptor knockout mouse.

The derangement of arterial endothelial cell morphology is a good indicator of a severe shock state. Because interleukin (IL)-1 has been implicated in this process, we examined the structural integrity of aortic endothelial cells in conjunction with serum IL-6 concentrations and nitric oxide levels, which are known to increase during endotoxemia in animals genetically devoid of the type 1 IL-1 receptor. Endotoxin (10 mg/kg Escherichia coli, injected intraperitoneally) (LD100) or saline vehicle was administered to adult male C57BL/129J wild-type control mice and C57BL/129J knockout mice possessing a homozygous deletion of the type 1 IL-1 receptor. The integrity of the aortic endothelium was determined by comparisons of ultrastructure. Mice injected with sterile vehicle showed normal endothelial ultrastructure with intact membranes. Wild-type and knockout control animals receiving saline vehicle showed a complete aortic endothelium (29.11 +/- .27 and 30.85 +/- .21 intact endothelial cells per millimeter of internal elastic lamina (IEL), respectively, p = N.S.). Endotoxin-treated wild-type animals showed extensive endothelial damage with most sections showing only denuded IEL on the luminal surface (1.83 +/- .38 cells/mm IEL, p < .001 vs. control). Knockout animals treated with endotoxin showed complete maintenance of endothelial structural integrity (34.08 +/- .57 cells/mm IEL, p < .001 vs. endotoxin-treated wild type) with ultrastructural morphology appearing identical to those given saline vehicle. Also, no apparent correlation was observed between serum IL-6 concentrations or serum nitric oxide levels and aortic endothelial damage. The maintenance of endothelial integrity in animals devoid of the IL-1 receptor confirms earlier observations of endothelial cell protection with IL-1 receptor antagonism and suggests that IL-1 contributes significantly to sepsis-induced endothelial damage.

The effects of epidural anesthesia on the neuroendocrine response to major surgical stress: a randomized prospective trial.

It has long been held that the acute-phase and neuroendocrine response to stress requires afferent neural input for its propagation. To further clarify the role of afferent neural impulses in this process and to determine the ability of epidural anesthesia to attenuate the normal perioperative stress response, 39 patients undergoing uncomplicated abdominal aortic replacement were randomized to receive either general anesthesia with postoperative patient-controlled intravenous morphine (n = 19) or combined regional/general anesthesia with intraoperative epidural catheter anesthesia using Bupivacaine to the T4 dermatome level followed by postoperative epidural morphine (n = 20). The stress response was quantitated by blinded measurement of baseline and postoperative (0, 12, 24, 48, and 72 hours) serum cortisol, epinephrine norepinephrine, total catecholamines, interleukin (IL)-1beta, IL-6, tumor necrosis factor (TNF)-alpha, and C-reactive protein (CRP). Total operative time (4.2 +/- 0.3 vs 4.3 +/- 0.4 hours), 72-hour fluid requirement (7.0 +/- 0.6 vs 6.8 +/- 0.71 mL), and length of hospitalization (7.8 +/- 1.4 vs 8.1 +/- 1.2 days) were not different between groups. All patients showed a significant increase in cortisol, epinephrine, norepinephrine, total catecholamines, CRP, and IL-6 in the postoperative period (P < 0.05). IL-1beta and TNF-alpha were less predictable and undetectable in most patients. There was no difference in any of the stress response indices between those patients receiving patient-controlled or epidural catheter anesthesia. In fact, the only parameter that was predictive of increased activation of the stress response was the length of operation, irrespective of anesthetic method. Those patients with operative times greater than 5 hours (n = 10) developed significantly higher CRP, IL-1beta, IL-6, and TNF-alpha levels (P < 0.05) at 12 and 24 hours postoperatively than those with total operative times less than 4 hours (n = 16). The neuroendocrine response to major surgical stress is propagated normally despite epidural blockade and is intensified with prolonged operative times. The inflammatory cytokines appear to play a major role in this process.

Does the direction of portal blood flow determine outcome with small-diameter prosthetic H-graft portacaval shunt?

BACKGROUND: Partial portal decompression, as attained by small-diameter prosthetic H-graft portacaval shunting, continues to gain popularity because of favorable outcomes. This study was undertaken to determine whether the direction of preshunt or postshunt portal blood flow or reversal in the direction of portal flow occurred with shunting effect outcome after small-diameter prosthetic H-graft portacaval shunt. METHODS: In 56 consecutive patients the direction of portal flow was determined before and after shunting. The direction of portal blood flow before and after shunting and changes in the direction of portal flow that occur with shunting were correlated with 30-day and 1-year survival, as well as with the rate of postshunt encephalopathy. RESULTS: Portal pressures significantly decreased in all with shunting. Whether or not stratified by Child's classification, neither the preshunt nor postshunt direction of portal flow affected 30-day or 1-year survival or incidence of encephalopathy. Eleven patients (significant at p < 0.001, fisher's exact test) had reversal of portal blood flow with shunting without an increase in 30-day or 1-year survival or incidence of encephalopathy. CONCLUSIONS: Small-diameter prosthetic H-graft portacaval shunts significantly reduce portal pressure and lead to reversal of portal flow in significant numbers. Significant hepatic dysfunction is uncommon after shunting. Neither the direction of preshunt or postshunt portal blood flow nor the reversal of portal blood flow with shunting has an effect on clinical outcome after small-diameter prosthetic H-graft portacaval shunt.

Specific changes in the pancreatic expression of the interleukin 1 family of genes during experimental acute pancreatitis.

Interleukin 1beta (IL-1beta) is produced in large amounts during acute pancreatitis and is believed to play a primary role in determining pancreatitis severity and the degree of pancreatic tissue destruction. This study was undertaken to characterize intrapancreatic production of IL-1beta and the remainder of the IL-1 family of genes during sterile acute pancreatitis. Moderate or severe necrotizing pancreatitis was induced by the intraperitoneal injection of a cholecystokinin analogue or the feeding of a choline deficient diet, respectively. Animals were killed during the progression of pancreatitis with severity scored by histological grading and serum amylase concentration. The expression of IL-1beta, IL-1 Receptor 1 (IL-1R1), Il-1R2, IL-1R antagonist (IL-1Ra), and ICE mRNA within the pancreas was examined by quantitative differential RT-PCR. Corresponding intrapancreatic and serum proteins were measured by enzyme-linked immunosorbent assay (ELISA). There was constitutive expression of pancreatic IL-1R1, IL-1R2, IL-1Ra, and ICE but not IL-1beta. As pancreatitis developed, mRNA for IL-1beta, IL-1Ra, and ICE increased in parallel with the degree of pancreatitis severity (all P<0.001 vs baseline) while mRNA for both receptors remained stable (P=NS). Intrapancreatic and systemic IL-1beta and IL-1Ra protein also increased as pancreatitis developed (both P<0.001) with tissue levels being continuously greater than serum. This study demonstrated that sterile, endotoxin-free acute pancreatitis induces the upregulation of specific members of the IL-1 family of genes including production of large amounts of IL-1beta and its receptor antagonist within the pancreatic parenchyma. These changes are indicative of pancreatitis severity and are not model dependent.

Differential sensitivity to Escherichia coli infection in mice lacking tumor necrosis factor p55 or interleukin-1 p80 receptors.

OBJECTIVE: To determine the effect of targeted disruption of the cellular receptors of either tumor necrosis factor alpha (TNF-alpha) or interleukin-1 beta (IL-1 beta) during experimental gram-negative bacterial infection and endotoxemia. DESIGN: Transgenic (knockout [KO]) mice deficient in either the p55 TNF receptor (TNF RI) or the p80 IL-1 receptor (IL-1 RI) were challenged with intravenous lipopolysaccharide (endotoxin) or intraperitoneal live Escherichia coli 0111:B4. Mortality was assessed daily for 7 days. Serum endotoxin levels and quantitative blood cultures were monitored at multiple times during infection. SETTING: Surgical infectious disease research laboratory. MAIN OUTCOME MEASURES: Mortality, results of quantitative blood cultures, and serum endotoxin levels. RESULTS: Both TNF and IL-1 RI KO mice were resistant to endotoxin challenge (0% mortality for both groups) compared with control mice (100% mortality [P < .01]). In contrast, only the IL-1 RI KO mice were resistant to infection caused by viable gram-negative bacteria (43% mortality) compared with control mice (100% mortality [P < .01]). Infection led to 100% mortality in TNF RI KO mice. The IL-1 RI KO mice exhibited less bacteremia and diminished endotoxemia compared with control and TNF RI KO mice 18 and 24 hours after infection. CONCLUSION: The absence of either the TNF or the IL-1 RI receptor prevents cellular activation by each respective cytokine. Absence confers protection against intravenous endotoxin, which stimulates massive rapid release of cytokines into the systemic circulation. However, bacterial infection within the peritoneal cavity is known to cause more delayed cytokine release, and cytokines may act at the site of infection to enhance host defenses. We believe that IL-1 signaling may be more critical in provoking lethal systemic toxic effects than TNF signaling. However, TNF signaling may be an important component of host defense enhancement at the local site of infection.