Emma Folch

Role of P-Selectin and ICAM-1 in Pancreatitis-Induced Lung Inflammation in Rats: Significance of Oxidative Stress

OBJECTIVE To investigate the role of P-selectin and intercellular adhesion molecule-1 (ICAM-1) in the pathogenesis of lung injury associated with pancreatitis, and the relation between xanthine oxidase-derived oxidants and expression of these adhesion molecules. SUMMARY BACKGROUND DATA In acute pancreatitis, acute respiratory distress syndrome occurs in the early stages of disease. This process is mediated by neutrophil infiltration. METHODS Pancreatitis was induced in rats by intraductal administration of 5% sodium taurocholate. ICAM-1 and P-selectin expression was measured using radiolabeled monoclonal antibodies. Neutrophil infiltration and plasma levels of xanthine oxidase were also evaluated. RESULTS Pancreatitis induces increases in P-selectin expression in lung, whereas ICAM-1 is unchanged from baseline levels. Immunoneutralization of either P-selectin or ICAM-1 prevents the infiltration of neutrophils into the lung. Xanthine and xanthine oxidase activity were increased after induction of pancreatitis. Xanthine oxidase inhibition prevents the upregulation of P-selectin in lung and neutrophil infiltration. CONCLUSIONS During acute pancreatitis, P-selectin is upregulated in the pulmonary endothelium and is a key determinant of leukocyte recruitment. Constitutive ICAM-1 is also involved in the process of cell infiltration into the lung. The increased expression of P-selectin appears to be triggered by a mechanism dependent on free radicals generated by xanthine oxidase released by the damaged pancreas.

Free radicals generated by xanthine oxidase mediate pancreatitis-associated organ failure.

In the present study we evaluate the possibilitythat xanthine oxidase released by damaged pancreas couldact as a source of oxidative damage in systemic tissuesduring the early stages of acute pancreatitis. This was accomplished by evaluating the effectsof xanthine oxidase inhibition with oxypurinol infusedinto the portal vein. Under these conditions, weinhibited the enzyme before it reached the liver and other distant organs, without inducing changesin the severity of pancreatic damage. Results indicatethat pancreatitis parallels increases in xanthineoxidase activity in plasma. Superoxide radicalsgenerated by this enzyme appears to be involved in thedecrease of reduced glutathione levels in the plasma andliver. In addition, xanthine oxidase inhibition preventsthe infiltration of neutrophils into the lungs. We conclude that oxygen free radicals generatedby xanthine and xanthine oxidase released to thebloodstream are involved in the systemic organ failureassociated with acute pancreatitis.

Leukotriene Generation and Neutrophil Infiltration After Experimental Acute Pancreatitis

The role of 5-lipoxygenase metabolites of arachidonic acid in the inflammatory response associated with experimental acute pancreatitis has been evaluated. For this purpose, an experimental necrohemorrhagic pancreatitis was induced in rats by intraductal administration of 5% sodium taurocholate. Neutrophil infiltration was detected in pancreas at 1 and 3 h after the induction of pancreatitis. This was concomitant with increased levels of leukotriene B4 and peptide leukotrienes (C4, D4 and E4). In lung, similar increases in neutrophil infiltration were detected but only 3 h after acute pancreatitis induction, and no changes in leukotriene B4 nor peptide leukotrienes were apparent at this time. These results suggest that after induction of acute pancreatitis, 5-lipoxygenase metabolites could play a role in the inflammatory response in the pancreas, but they are not involved in the inflammatory response in lung.

P-selectin expression and Kupffer cell activation in rat acute pancreatitis

This work studied the activation of hepatic macrophages during acute pancreatitis and the involvement of these cells in the lung inflammatory response. Pancreatitis was induced in Wistar rats by intraductal administration of 5% sodium taurocholate. Three hours after pancreatitis induction, the degree of pulmonary inflammation, TNF-α levels, and P-selectin expression were evaluated. The generation of TNF-α by Kupffer cells was also measured. Pancreatitis increases the serum concentration of TNF-α, neutrophil infiltration, and P-selectin expression in pancreas and lung. In addition, Kupffer cells generate increased levels of TNF-α. When Kupffer cells were inhibited, the increase in serum TNF-α levels and the infiltration of neutrophils in the lung were prevented, but P-selectin expression remained unmodified. We conclude that pulmonary inflammation induced by acute pancreatitis is mediated by Kupffer cell activation and that pancreatitis induces the expression of P-selectin on pulmonary endothelial cells but this effect is not mediated by Kupffer cells.

H2O2 and PARS mediate lung P-selectin upregulation in acute pancreatitis

P-selectin and circulating xanthine oxidase are involved in the process of neutrophil infiltration into the lung associated with acute pancreatitis. This study investigated the mediators that trigger the upregulation of P-selectin in this process. Pancreatitis was induced in rats by intraductal administration of 5% sodium taurocholate. P-selectin expression was measured using radiolabeled antibodies. Neutrophil infiltration and PAF levels were also evaluated. The role of superoxide radical, H(2)O(2), or the enzyme poly (ADP-ribose) synthetase (PARS) on these processes was determined in groups of animals treated with the corresponding inhibitors. Pancreatitis was associated with an increase in P-selectin expression in the lung. Inhibition of PARS or H(2)O(2) abrogated P-selectin upregulation, PAF generation, and neutrophil recruitment. Superoxide dismutation prevented neutrophil recruitment and PAF generation, but had no effect on P-selectin expression. We conclude that during acute pancreatitis, upregulation of P-selectin in the pulmonary endothelium is triggered by H(2)O(2) and PARS activity.

Soluble receptors released during acute pancreatitis interfere with the detection of tumor necrosis factor-α

ObjectiveTo evaluate the interfering effect of tumor necrosis factor-&agr; soluble receptor when measuring circulating concentrations of tumor necrosis factor-&agr; in an experimental model of acute pancreatitis. DesignRandomized, controlled trial. SettingExperimental laboratory. SubjectsMale Wistar rats. InterventionsAcute pancreatitis was induced by intraductal administration of 5% sodium taurocholate. Saline was administered in a control group. Serums were overloaded with known amounts of tumor necrosis factor-&agr; or macrophage inflammatory protein-2. Measurements and Main Results Three hours after induction, serum concentrations of free tumor necrosis factor-&agr;, total tumor necrosis factor-&agr;, and soluble receptor of tumor necrosis factor-&agr; were measured. No detectable concentrations of free tumor necrosis factor-&agr; were found in any experimental group. By contrast, significant increases in total tumor necrosis factor-&agr; and soluble receptor of tumor necrosis factor-&agr; were found after induction of pancreatitis. Overloading of serum with tumor necrosis factor-&agr; resulted in detection of 50% of the expected concentrations of free tumor necrosis factor-&agr; from control animals and only of 5% from the pancreatitis group. Overloading the serum with macrophage inflammatory protein-2 resulted in a detection of 100% of the expected concentrations in both control and treated animals. ConclusionCirculating soluble receptor of tumor necrosis factor-&agr; could interfere with the detection of tumor necrosis factor-&agr; in some pathologies, such as pancreatitis, that are associated with increases in soluble receptor of tumor necrosis factor-&agr;.

Absorption and effects of 3-(N-phenylamino)-1,2-propanediol esters in relation to Toxic Oil Syndrome

Toxic Oil Syndrome (TOS) was an epidemic disease related to the consumption of rapessed oil denatured with aniline that made its sudden appearance in Spain in 1981. The fatty acid esters of 3-(N-phenylamino)-1,2-propanediol (PAP), which is a chemical class of by-products resulting from the reaction of aniline with oil components, have shown a strong association with TOS-related oils. These compounds also show some structural similarities to platelet-activating factor (PAE). In search of a toxic agent that could explain the widespread systemic effects observed in TOS patients, we investigated the intestinal absorption and biotransformation of the different PAP esters found in TOS-related oil samples and the possible pathophysiological effect of these mediators and their metabolic products if acting as PAF analogs. Results indicate that PAP esters are absorbed in the gastrointestinal tract and are distributed and stored in different organs, particularly in the liver and brown adipose tissue. PAP in these organs showed different patterns of fatty acids, indicating the ability of the gastrointestinal tract to modify the fatty acid composition of the parent PAP. Thus, the fatty acid profile of the PAP esters found in intestine appears to be related to the type of oil used as vehicle. Some of these PAP esters, when a long acyl chain was present in the sn-1 position of the molecule, showed an inhibitory effect on the PAF synthesis. This is an important observation in line with the systemic nature of the disease.

Effect of peritoneal lavage and lymph ligature on systemic complications of experimental acute pancreatitis

We studied the involvement of ascitic fluid on the systemic effects of experimental acute pancreatitis. This has been achieved by comparing the effectiveness of either peritoneal lavage or lymphatic ligature on preventing changes in systemic vascular permeability. Three hours after induction of pancreatitis, we found increases in vascular permeability in the pancreas, lung, and intestine. Both peritoneal lavage and lymphatic ligature were able to prevent the changes observed in the lung and intestine and the increases on plasma levels of lipase and amylase, suggesting a similar involvement for lymphatic draining and peritoneal absorption pathways. In addition, we evaluated the effect of intraperitoneal deposition into health rats of pancreatitis-associated ascitic fluid collected from rats with experimental acute pancreatitis. A significant increase in plasma amylase and lipase levels could be observed but no changes in vascular permeability were found. Altogether, these results indicate that transperitoneal absorption of toxic mediators from the ascitic fluid is not enough to explain the systemic damage induced by acute pancreatitis.

GSK3β and VDAC Involvement in ER Stress and Apoptosis Modulation during Orthotopic Liver Transplantation

We investigated the involvement of glycogen synthase kinase-3β (GSK3β) and the voltage-dependent anion channel (VDAC) in livers subjected to cold ischemia–reperfusion injury (I/R) associated with orthotopic liver transplantation (OLT). Rat livers were preserved in University of Wisconsin (UW) and Institute Georges Lopez (IGL-1) solution, the latter enriched or not with trimetazidine, and then subjected to OLT. Transaminase (ALT) and HMGB1 protein levels, glutamate dehydrogenase (GLDH), and oxidative stress (MDA) were measured. The AKT protein kinase and its direct substrates, GSK3β and VDAC, as well as caspases 3, 9, and cytochrome C and reticulum endoplasmic stress-related proteins (GRP78, pPERK, ATF4, and CHOP), were determined by Western blot. IGL-1+TMZ significantly reduced liver injury. We also observed a significant phosphorylation of AKT, which in turn induced the phosphorylation and inhibition of GSK3β. In addition, TMZ protected the mitochondria since, in comparison with IGL-1 alone, we found reductions in VDAC phosphorylation, apoptosis, and GLDH release. All these results were correlated with decreased ER stress. Addition of TMZ to IGL-1 solution increased the tolerance of the liver graft to I/R injury through inhibition of GSK3β and VDAC, contributing to ER stress reduction and cell death prevention.