Search

Antimicrobial peptides (AMPs) play essential roles in maintaining intestinal health and have been suggested as possible therapeutic strategies against inflammatory bowel disease (IBD). However, the instability of AMPs in the process of transmission in vivolimits their application in the treatment of IBD. In this study, we constructed the mBD14-producing Lactococcus lactisNZ9000 (L. lactis/mBD14) to achieve enteric delivery of mBD14 and evaluated its protective effect on dextran sodium sulfate (DSS)-induced colitis. Mice treated with L. lactis/mBD14 exhibited milder symptoms of colitis (P< 0.01). Additionally, L. lactis/mBD14 treatment reversed DSS-induced epithelial dysfunction and reduced the production of pro-inflammatory cytokines in colon (P< 0.01). Mechanistically, L. lactis/mBD14 significantly inhibited NOD-like receptor pyrin domain containing three inflammasome-mediated pro-inflammatory response (P< 0.05) and regulated microbiota homeostasis by promoting the abundance of probiotic bacteria Akkermansia muciniphilaand Faecalibacterium prausnitziiand decreasing the pathogenic Escherichia coli(P< 0.01). Taken together, this study demonstrates the protective effect of L. lactis/mBD14 in DSS-induced colitis, and suggests that oral administration of L. lactis/mBD14 may represent a potential therapeutic strategy for IBD.

Introduction:The view of hydrogen sulfide has changed from a toxic by-product to a crucial signaling molecule, with enormous potential as a pharmacological target for diseases ranging from heart disease to sepsis. Despite this progression of ideas, there is still a large amount that is unknown about this gaseous signaling molecule. Hydrogen sulfide has been implicated as a tissue protectant in many pathological conditions, the mechanisms of tissue protection is a point of controversy, particularly distinguishing the direct actions from the indirect downstream effects of hydrogen sulfide. This point of controversy is particularly pertinent in inflammation research.Areas covered:Current research into the pathways in which hydrogen sulfide can act as a pro-inflammatory molecule and as an anti-inflammatory molecule.Expert opinion:How controversies regarding hydrogen sulfide may have occurred is discussed. Addressed are the direct and indirect pathways of hydrogen sulfide on inflammation, the effects of different concentrations of hydrogen sulfide and how the effects of hydrogen sulfide on the immune system vary with different delivery mechanisms. Furthermore, there is a discussion on what key gaps exist in current knowledge and must be addressed before hydrogen sulfide can be considered a valid pharmacological target.

This study aimed to determine the effect of hydrogen sulfide (H2S) on Toll-like receptor 4 (TLR4)-mediated innate immune signaling in acute pancreatitis (AP) via substance P.

Hydrogen sulfide (H2S), the gaseous mediator produced by various cells in our body, was recently discovered to play a major role in human physiology despite its toxic nature known for centuries. In addition to its pathophysiological relevance in cardiovascular and neuronal disorders, there is considerable interest in the significance of H2S in inflammation. A number of preclinical studies in our laboratory as well as by others, using H2S donors and inhibitors of its endogenous synthesis, have provided evidence for both pro- and anti-inflammatory character of H2S. But so far, there is a significant lack of support from relevant clinical studies. One of the major contentious issues being variable dose and sampling time, controversies exist on the precise friend or foe nature of this gaseous transmitter. However, it is well accepted that once a clearer picture of the whole story of H2S in inflammation emerges, potential for therapeutic manipulations in this field are immense. This review focuses on the intriguing effects of H2S in some of the inflammatory conditions such as acute pancreatitis, sepsis, burn injuries and local inflammation of the joints. Active research projects have been undertaken to elucidate the mechanisms of action of H2S in inflammation, including neurogenic inflammation and interaction with other biological mediators and pathways. The early and fragmentary evidence obtained holds promise for a successful drug intervention for these inflammatory diseases.

We previously reported a high incidence of alcohol-related acute pancreatitis (AP) in Goa, India, where country-made alcoholic products are consumed in addition to the commercially available alcoholic products. We aimed to analyze the composition of these country-made alcoholic products consumed by a population with a high incidence of alcohol-related AP.

The neuropeptide substance P (SP) has emerged to be an important proinflammatory mediator in acute pancreatitis (AP). The presence of substance P and its receptor, neurokinin-1 receptor (NK1R) has been shown in the pancreas and the pancreatic acinar cells. In this study, we investigated the unexplored mechanisms that mediate SP and NK1R expression using an in vitro AP model. Pancreatic acinar cells were obtained from pancreas of male Swiss mice. Isolated cells were treated with caerulein to mimic secretagogue pancreatitis. A concentration-dependent study that subjected the cells to 60 min of stimulation by caerulein showed that SP and the transcript from its gene preprotachykinin-A (PPT-A), and NK1R were up-regulated at a supraphysiological concentration of 10(-7) M. A concentration-dependent study on intracellular kinases, extracellular signal-regulated kinase (ERK1/2), and c-Jun N-terminal kinase (JNK) and also transcription factors nuclear factor-kappaB (NF-kappaB) and activator protein-1 (AP-1) showed that they were activated when the caerulein concentration was 10(-7) M. Inhibition of JNK reversed the up-regulation of PPT-A, SP, and NK1R. However, inhibition of ERK1/2 reversed the up-regulation of NK1R but not of PPT-A and SP. Furthermore, we found that specific ERK1/2 and JNK inhibitors reduce NF-kappaB and AP-1 activity. Taken together, our results suggest that supraphysiological concentrations of caerulein up-regulate the expression of SP and NK1R in pancreatic acinar cells, and the signaling molecules that are involved in this up-regulation include ERK1/2, JNK, NF-kappaB, and AP-1.

Acute pancreatitis (AP) and sepsis are inflammatory disorder varying in magnitude of response to infection or inflammatory stimuli. The specific role of various causative factors in AP, septic shock, current pharmacological treatments, animal models, role of infiltrating cells and novel molecules that play an important role in the disease progression to sepsis are explored. AP is an inflammatory disease of the pancreas. Over the years accumulating evidence suggests numerous molecules as key regulators of the inflammatory signaling cascade such as selectins, chemokine signaling and expression of intergrins on leukocytes facilitate adhesion to vessel wall. Inhibition of any of these molecules has proven to be effective in animal models of AP. Recently, the biochemical role of hydrogen sulfide (H2S) and substance P in caerulein induced AP and in cecal ligation and puncture induced sepsis and their role in the pathogenesis of the disease have highlighted the importance of novel molecules as therapeutic targets in addition to the known proinflammatory molecules, cytokines and chemoattractant chemokines and their receptors upregulated in AP and sepsis. This review aims to give an overview of the multifaceted complex interactions in a prearranged fashion and their functional role in the inflammatory process that afflict AP and sepsis. The interlinking molecules in AP and sepsis emphasize the similarities in the inflammatory response and the importance of pharmacological agents that reduce or inhibit the progression to chronic stage.

We have shown earlier that mouse pancreatic acinar cells produce hydrogen sulfide (H(2)S) and play a role in the pathogenesis of acute pancreatitis. It is noteworthy that recent evidence indicates that H(2)S has anti-inflammatory effects. To date, the mechanism by which H(2)S directly reduces inflammation has not been elucidated. In the present study, we hypothesized that H(2)S inhibits the production of proinflammatory cytokines by activation of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway. Pancreatic acinar cells were treated with the H(2)S donor, sodium hydrogen sulfide (NaHS) (5, 10, and 30 microM). To better understand the effect of H(2)S in inflammation, pancreatic acinar cells were stimulated with caerulein after the addition of NaHS (5, 10, and 30 microM). We observed that H(2)S at the 5 microM concentration down-regulates the activation of NF-kappaB and degradation of IkappaB alpha. However, H(2)S (5 microM) activates PI3K as reflected by AKT phosphorylation. We found that H(2)S-mediated activation of PI3K in caerulein-treated acinar cells correlated with the down-regulation of extracellular signal-regulated kinase 1/2 phosphorylation, whereas phosphorylation of p38 and c-Jun NH(2)-terminal kinase and mitogen-activated protein kinases was unchanged. The PI3K inhibitor LY294002 [2-(4-morpholinyl)-8-phenyl-1(4H)-benzopyran-4-one hydrochloride] abolished the H(2)S-mediated activation of AKT and increases tumor necrosis factor alpha and interleukin 1beta levels in caerulein-treated acinar cells. These findings indicate that the phosphatidylinositol 3-kinase plays a negative role in NaHS-treated pancreatic acinar cells and suggest a role for H(2)S in the PI3K/AKT pathway in acute pancreatitis.

Substance P is known to play a key role in the pathogenesis of acute pancreatitis. Src family kinases (SFKs) are known to be involved in cytokine signaling. However, the involvement of SFKs in substance P-induced chemokine production and its role in acute pancreatitis have not been investigated yet. To that end, we have used primary preparations of mouse pancreatic acinar cells as our model to show that substance P/neurokinin 1 receptor (NK1R) induced activation of SFKs. SFKs mediated the activation of mitogen-activated protein kinases [extracellular signal-regulated kinase (ERK), c-Jun NH(2)-terminal kinase (JNK)], transcription factors [signal transducer and activator of transcription (STAT) 3, nuclear factor (NF) kappaB, activator protein-1 (AP-1)], and production of chemokines in pancreatic acinar cells. We further tested the significance of the SFK signaling pathway in acute pancreatitis. Our results show, for the first time, that treatment of mice with the potent and selective SFK inhibitor PP2 [4-amino-5-(4-chlorophenyl)-7-(t-butyl) pyrazolo [3,4-D] pyrimidine], but not its negative inhibitor PP3 (4-amino-7-phenylpyrazol [3,4-D] pyrimidine), reduced the severity of pancreatitis. This was proven by significant attenuation of hyperamylasemia, pancreatic myeloperoxidase activity, chemokines, and water content. Histological evidence of diminished pancreatic injury also confirmed the protective effect of the inhibition of SFKs. Moreover, treatment with the substance P receptor antagonist CP96345 [(2S,3S)-cis-2-(diphenylmethyl)-N-((2-methoxyphenyl)-methyl)-1-azabicyclo(2.2.2.)-octan-3-amine] attenuated acute pancreatitis-induced activation of SFKs, ERK, JNK, STAT3, NFkappaB, and AP-1. The proposed signaling pathway through which substance P mediates acute pancreatitis is through substance P/NK1R-SFKs-(ERK, JNK)-(STAT3, NFkappaB, AP-1) chemokines. In light of our study, we propose that drugs targeting the substance P-mediated signaling pathways could prove beneficial in improving treatment efficacy in acute pancreatitis.

Background/Aims:The role of nitric oxide (NO) has been increasingly implicated in the pathophysiology of acute pancreatitis (AP). Studies have shown increased NO production in AP although not all are agreeable on whether NO is beneficial or detrimental in AP. This study aims to profile NO production and NO synthase (NOS) expression in the pancreas and lungs in the progression of AP in mice to gain insights to the role played by different NOS isoforms. Methods:AP was induced in mice by hourly administration of cerulein. NO production was determined by measuring the total nitrite and nitrate (NOx) content while NOS expression was measured by Western blot. Results:Pancreatic NO production increased sharply and was sustained throughout AP. iNOS expression was greatly increased while eNOS was downregulated at the later stages. In the lungs, there was an unexpected early increase in the constitutive NOS expression; however iNOS was also significantly overexpressed at the later time point along with a significant increase in NO. Acinar cells were found to overproduce NO in response to cerulein hyperStimulation with iNOS again being the major contributor. Conclusion:These data show that NO production and NOS expression are differentially regulated temporally and in magnitude in the pancreas and lungs in response to cerulein hyperstimulation which suggests differing roles for each NOS isoform.

Fractalkine (FTK) is a unique member of the CX3C chemokine family by acting through the CX3CR1 receptor. Membrane-bound FTK acts like an adhesion molecule, whereas soluble FTK (sFTK) acts as a classic chemokine ligand. Whether this chemokine plays a role in sepsis is still not clear. Using a mouse model of cecal ligation and puncture (CLP)-induced sepsis, we found that FTK levels were elevated in plasma 24 h after CLP. Reverse transcription-polymerase chain reaction results showed that FTK messenger RNA levels were upregulated, whereas CX3CR1 messenger RNA levels were downregulated in lungs after CLP procedure. To study the role of FTK in lung injury during sepsis, we injected exogenous sFTK into the mice before the CLP procedure. We found that plasma FTK levels were further elevated by sFTK. Mice that were injected with FTK had a lower myeloperoxidase activity in lungs compared with the CLP group. Furthermore, macrophage inflammatory protein 2, IL-1, and IL-6 levels in lungs were reduced after the injection of FTK. Treatment with sFTK also attenuated lung morphological changes in histological sections. To find out whether sFTK had an effect on leukocyte rolling and adherence, intravital microscope was used. Results showed that sFTK significantly attenuated leukocyte adhesion but had little effect on leukocyte rolling in mesenteric microcirculation. Taken together, our findings suggest that FTK may be a novel chemokine that modulates neutrophil infiltration and chemokine and cytokine production during sepsis.

AbstractBackground/Aims:The role of nitric oxide (NO) has been increasingly implicated in the pathophysiology of acute pancreatitis (AP). Studies have shown increased NO production in AP although not all are agreeable on whether NO is beneficial or detrimental in AP. This study aims to profile NO production and NO synthase (NOS) expression in the pancreas and lungs in the progression of AP in mice to gain insights to the role played by different NOS isoforms. Methods:AP was induced in mice by hourly administration of cerulein. NO production was determined by measuring the total nitrite and nitrate (NOx) content while NOS expression was measured by Western blot. Results:Pancreatic NO production increased sharply and was sustained throughout AP. iNOS expression was greatly increased while eNOS was downregulated at the later stages. In the lungs, there was an unexpected early increase in the constitutive NOS expression; however iNOS was also significantly overexpressed at the later time point along with a significant increase in NO. Acinar cells were found to overproduce NO in response to cerulein hyperstimulation with iNOS again being the major contributor. Conclusion:These data show that NO production and NOS expression are differentially regulated temporally and in magnitude in the pancreas and lungs in response to cerulein hyperstimulation which suggests differing roles for each NOS isoform.Copyright © 2008 S. Karger AG, Basel and IAP

The present study investigated whether chemokines are involved in hydrogen sulfide (H2S)-associated pathogenesis of acute pancreatitis and associated lung injury.

This study aimed to test these hypotheses: cystathionine gamma-lyase (CSE) is expressed in a human artery, it generates hydrogen sulfide (H(2)S), and H(2)S relaxes a human artery. H(2)S is produced endogenously in rat arteries from cysteine by CSE. Endogenously produced H(2)S dilates rat resistance arteries. Although CSE is expressed in rat arteries, its presence in human blood vessels has not been described. In this study, we showed that both CSE mRNA, determined by reverse transcription-polymerase chain reaction, and CSE protein, determined by Western blotting, apparently occur in the human internal mammary artery (internal thoracic artery). Artery homogenates converted cysteine to H(2)S, and the H(2)S production was inhibited by dl-propargylglycine, an inhibitor of CSE. We also showed that H(2)S relaxes phenylephrine-precontracted human internal mammary artery at higher concentrations but produces contraction at low concentrations. The latter contractions are stronger in acetylcholine-prerelaxed arteries, suggesting inhibition of nitric oxide action. The relaxation is partially blocked by glibenclamide, an inhibitor of K(ATP) channels. The present results indicate that CSE protein is expressed in human arteries, that human arteries synthesize H(2)S, and that higher concentrations of H(2)S relax human arteries, in part by opening K(ATP) channels. Low concentrations of H(2)S contract the human internal mammary artery, possibly by reacting with nitric oxide to form an inactive nitrosothiol. The possibility that CSE, and the H(2)S it generates, together play a physiological role in regulating the diameter of arteries in humans, as has been demonstrated in rats, should be considered.

Endotoxemia is a life‐threatening, inflammatory condition that involves multiple organ injury and dysfunction. Preprotachykinin‐A (PPT‐A) gene products, substance P (SP), and neurokinin‐A have been shown to play an important role in neurogenic inflammation. To investigate the role of PPT‐A gene products on multiple organ injury in LPS‐induced endotoxemia, endotoxemia was induced by LPS administration (10 mg/kg, i.p.) in PPT‐A gene‐deficient mice (PPTA−/−) and the wild‐type (WT) control mice (PPT‐A+/+). I.p. administration of LPS to WT mice caused a significant increase in circulating levels of SP as well as in liver, lung, and kidney. PPT‐A gene deletion significantly protected against liver, pulmonary, and renal injury following LPS‐induced endotoxemia, as evidenced by tissue myeloperoxidase activities, plasma alanine aminotransferase, aspartate aminotransferase levels, and histological examination. Furthermore, PPT‐A−/−mice had significantly attenuated chemokines, proinflammatory cytokines, and adhesion molecule levels in the liver, lung, and kidney. These results show that PPT‐A gene products are critical proinflammatory mediators in endotoxemia and the associated multiple organ injury. In addition, the data suggest that deletion of the PPT‐A gene protected mice against organ damage in endotoxemia by disruption in neutrophil recruitment.

Impaired lung function in severe acute pancreatitis is the primary cause of morbidity and mortality in this condition. Hydrogen sulfide (H2S) is a naturally occurring gas that has been shown to be a potent vasodilator. Diclofenac is a nonsteroidal anti-inflammatory drug and has been shown to have anti-inflammatory, analgesic, and antipyretic activity. ACS15 is an H2S-releasing derivative of diclofenac. Little is known about its effectiveness as an anti-inflammatory drug. In this report, we describe the effect of diclofenac and its H2S-releasing derivative on acute pancreatitis and associated lung injury in the mouse. Acute pancreatitis was induced in mice by hourly i.p. injections of cerulein. Diclofenac and ACS15 were administered either 1 hour before or 1 hour after starting cerulein injections, and the severity of acute pancreatitis and associated lung injury was assessed. The severity of acute pancreatitis was determined by hyperamylasemia, neutrophil sequestration in the pancreas (pancreatic myeloperoxidase activity), and pancreatic acinar cell injury/necrosis on histological examination of pancreas sections. The severity of acute pancreatitis-associated lung injury was assessed by neutrophil sequestration in the lungs (lung myeloperoxidase activity) and by histological examination of lung sections. ACS15, given prophylactically and therapeutically, significantly reduced lung inflammation without having any significant effect on pancreatic injury. These results suggest the usefulness of H2S-releasing nonsteroidal anti-inflammatory drugs as potential treatments for pancreatitis-associated lung injury.

Hydrogen sulfide (H2S) is recognized increasingly as a proinflammatory mediator in various inflammatory conditions. Here, we have investigated the role of H2S in regulating expression of some endothelial adhesion molecules and recruitment of leukocytes to inflamed sites in sepsis. Male Swiss mice were subjected to cecal ligation and puncture (CLP)‐induced sepsis and treated with saline (i.p.), DL‐propargylglycine (PAG; 50 mg/kg, i.p.), an inhibitor of H2S formation or NaHS (10 mg/kg, i.p.), an H2S donor. PAG was administered 1 h before or after the induction of sepsis, and NaHS was given at the same time of CLP. Using intravital microcopy, we found that in sepsis, prophylactic and therapeutic administration of PAG reduced leukocyte rolling and adherence significantly in mesenteric venules coupled with decreased mRNA and protein levels of adhesion molecules (ICAM‐1, P‐selectin, and E‐selectin) in lung and liver. In contrast, injection of NaHS up‐regulated leukocyte rolling and attachment significantly, as well as tissue levels of adhesion molecules in sepsis. Conversely, normal mice were given NaHS (10 mg/kg, i.p.) to induce lung inflammation, with or without NF‐κB inhibitor BAY 11‐7082 pretreatment. NaHS treatment enhanced the level of adhesion molecules and neutrophil infiltration in lung. These alterations were reversed by pretreatment with BAY 11‐7082. Moreover, expression of CXCR2 in neutrophils obtained from H2S‐treated mice was up‐regulated significantly, leading to an obvious elevation in MIP‐2‐directed migration of neutrophils. Therefore, H2S acts as an important endogenous regulator of leukocyte activation and trafficking during an inflammatory response.

Earlier work from our laboratory has suggested a role for the neuropeptide substance P (SP) in inducing lung injury in sepsis. In that study, mice lacking the preprotachykinin‐A gene, which encodes for SP, were protected against lung injury in sepsis. To further substantiate the role of SP in sepsis and to study its mchanism, we have evaluated the effect of SR140333, a SP receptor antagonist, on lung injury in sepsis, which was induced in male Swiss mice by cecal ligation and puncture (CLP). Sham‐operated animals received the same surgical procedure, except CLP. Vehicle or SR140333 (1 mg/kg, s.c.) was administered to CLP mice 30 min before or 1 h after the CLP. Eight hours after surgery, lung tissue was collected and analyzed for myeloperoxidase (MPO) activity, chemokines, cytokines, and adhesion molecules. The CLP procedure alone caused a significant increase in the lung levels of MIP‐2, MCP‐1, IL‐1β, IL‐6, ICAM‐1, E‐ and P‐selectin, and MPO activity when compared with sham‐operated mice. SR140333 injected 30 min before or 1 h after CLP significantly attenuated the increased lung MPO activity and levels of MIP‐2, MCP‐1, IL‐1β, IL‐6, ICAM‐1, and E‐ and P‐selectin compared with CLP‐operated mice injected with the vehicle. Histological evaluation of the lung sections further supported the beneficial effect of SR140333 on lung inflammation. Therefore, SP receptor antagonism can be a potential therapeutic target in polymicrobial sepsis, and this effect is brought about via reduction in leukocyte recruitment.

Apoptosis is a teleologically beneficial form of cell death in acute pancreatitis. Our previous work has demonstrated that induction of pancreatic acinar cell apoptosis by crambene protects mice against acute pancreatitis. However, little is known about how the induction of apoptosis reduces the severity of acute pancreatitis. Because the clearance of apoptotic cells might suppress inflammation and critically regulate immune responses, we postulate that clearance of apoptotic cells stimulates an anti-inflammatory response, which has a protective action against acute pancreatitis. To test this hypothesis, induction of apoptosis in acute pancreatitis in vivoand co-cultures of peritoneal resident macrophages with apoptotic acinar cells in vitrowere used as experimental systems, testing expression of phagocytic receptors and levels of inflammatory mediators. Moreover, neutralizing anti-interleukin (IL)-10 monoclonal antibody (2.5 mg/kg) was used before the induction of apoptosis in acute pancreatitis, testing whether the protection from apoptosis induction would be removed. Our study showed that clearance of apoptotic acinar cells, which may occur essentially through the CD36-positive macrophage, stimulates the release of anti-inflammatory mediators like IL-10. IL-10 plays an important role in crambene-induced protection in acute pancreatitis. Thus, induction of pancreatic acinar cell apoptosis by crambene protects mice against acute pancreatitis via induction of anti-inflammatory pathways.

Hydrogen sulfide (H2S) is now considered an endogenous, gaseous mediator, which has been demonstrated to be involved in many inflammatory states. However, the mechanism of its proinflammatory function remains unknown. In the present study, we used IFN‐γ‐primed human monocyticcell line U937 to investigate the effects of H2S in vitro on monocytes. We found that treatment with the H2S donor, sodium hydrosulfide, led to significant increases in the mRNA expression and protein production of TNF‐α, IL‐1β, and IL‐6 in U937 cells. H2S‐triggered monocyte activation was confirmed further by the up‐regulation of CD11b expression on the cell surface. We also observed that H2S could induce a rapid degradation of IκBα and subsequent activation of NF‐κB p65, and this effect was attenuated by Bay 11‐7082, a specific inhibitor of NF‐κB. Furthermore, pretreatment of cells with Bay 11‐7082 substantially inhibited the secretion of TNF‐α, IL‐1β, and IL‐6 induced by H2S. We also found that H2S stimulated the phosphorylation and activation of ERK1/2, but not of p38 MAPK and JNK, and pretreatment with PD98059, a selective MEK1 antagonist, could inhibit H2S‐induced NF‐κB activation markedly. Together, our findings suggest for the first time that H2S stimulates the activation of human monocytes with the generation of proinflammatory cytokines, and this response is, at least partially, through the ERK‐NF‐κB signaling pathway.

Chemokines and their receptors play a key role in the pathogenesis of acute pancreatitis. BX471 is a potent nonpeptide CC chemokine receptor 1 antagonist in both human and mouse. The aim of the present study was to evaluate the effect of prophylactic and therapeutic treatment with BX471 on experimental acute pancreatitis in the mouse and to investigate the underlying mechanisms.

Impaired lung function in severe acute pancreatitis is the primary cause of morbidity and mortality in this condition. Flurbiprofen is a powerful nonsteroidal anti-inflammatory drug (NSAID). However, administration of this drug is associated with severe gastrointestinal side effects. The NO-releasing derivative of flubiprofen (nitroflurbiprofen, HCT1026) has recently been developed by the addition of a nitroxybutyl moiety to the flurbiprofen structure. This modification does not interfere with the anti-inflammatory activity of the drug but markedly reduces its ability to induce gastric injury. The effects of treatment with flurbiprofen and HCT1026 on the severity of pancreatitis and the associated lung injury were investigated in a mouse model. Acute pancreatitis was induced in mice by hourly intraperitoneal injections of cerulein. Flurbiprofen and HCT1026 were administered either 30 min before or 1 h after starting cerulein injections, and the severity of acute pancreatitis and associated lung injury were assessed. The severity of acute pancreatitis was determined by hyperamylasemia, neutrophil sequestration in the pancreas (pancreatic MPO activity), and pancreatic acinar cell injury/necrosis on histological examination of pancreas sections. The severity of acute pancreatitis-associated lung injury was assessed by neutrophil sequestration in the lungs (lung MPO activity) and by histological examination of lung sections. HCT1026 and flurbiprofen, given prophylactically as well as therapeutically, significantly reduced lung inflammation without having any significant effect on pancreatic injury. These results suggest the usefulness of flurbiprofen as well as HCT1026 as potential treatments for pancreatitis-associated lung injury.

Inflammatory mediators play a critical role in acute pancreatitis. The precise role played by members of the chemokine family remains unclear.

Inflammatory mediators play a key role in acute pancreatitis and the resultant multiple organ dysfunction syndrome, which is the primary cause of death in this condition. Recent studies have confirmed the critical role played by inflammatory mediators such as TNF-α, IL-1β, IL-6, IL-8, PAF, IL-10, C5a, ICAM-1, and substance P. The systemic effects of acute pancreatitis have many similarities to those of other conditions such as septicaemia, severe burns, and trauma. The delay between the onset of inflammation in the pancreas and the development of the systemic response makes acute pancreatitis an ideal experimental and clinical model with which to study the role of inflammatory mediators and to test novel therapies. Elucidation of the key mediators involved in the pathogenesis of acute pancreatitis will facilitate the development of clinically effective anti-inflammatory therapy. Copyright © 2000 John Wiley & Sons, Ltd.

Inflammatory mediators play a key role in acute pancreatitis and the resultant multiple organ dysfunction syndrome, which is the primary cause of death in this condition. Recent studies have confirmed the critical role played by inflammatory mediators such as TNF‐α, IL‐1β, IL‐6, IL‐8, PAF, IL‐10, C5a, ICAM‐1, and substance P. The systemic effects of acute pancreatitis have many similarities to those of other conditions such as septicaemia, severe burns, and trauma. The delay between the onset of inflammation in the pancreas and the development of the systemic response makes acute pancreatitis an ideal experimental and clinical model with which to study the role of inflammatory mediators and to test novel therapies. Elucidation of the key mediators involved in the pathogenesis of acute pancreatitis will facilitate the development of clinically effective anti‐inflammatory therapy. Copyright © 2000 John Wiley & Sons, Ltd.

Background & Aims:Intercellular adhesion molecule 1 (ICAM-1) and neutrophils play important roles in many inflammatory processes, but their importance in both acute pancreatitis and pancreatitis-associated lung injury has not been defined. Methods:To address this issue, mice that do not express ICAM-1 were used and depleted of neutrophils by administration of antineutrophil serum. Pancreatitis was induced by administering either supramaximal doses of the secretagogue cerulein or feeding a choline-deficient, ethionine-supplemented diet. The severity of pancreatitis was evaluated by quantitating serum amylase, pancreatic edema, acinar cell necrosis, and pancreas myeloperoxidase activity (i.e., neutrophil content). Lung injury was evaluated by quantitating lung myeloperoxidase activity and pulmonary microvascular permeability. ICAM-1 was quantitated by enzyme-linked immunosorbent assay and was localized by light-microscopic immunohistochemistry. Results:It was found that serum, pancreas, and lung ICAM-1 levels increase during pancreatitis. Both pancreatitis and the associated lung injury are blunted, but not completely prevented, in mice deficient in ICAM-1. Neutrophil depletion also reduces the severity of both pancreatitis and lung injury. However, the combination of neutrophil depletion with ICAM-1 deficiency does not reduce the severity of pancreatitis or lung injury to a greater extent than either neutrophil depletion or ICAM-1 deficiency alone. Neither pancreatitis nor pancreatitis-associated lung injury are completely prevented by ICAM-1 deficiency, neutrophil depletion, or combined ICAM-1 deficiency plus neutrophil depletion. Conclusions:The observations indicate that ICAM-1 plays an important, neutrophil-mediated, proinflammatory role in pancreatitis and pancreatitis-associated lung injury. The studies also indicate that ICAM-1 and neutrophil-independent events also contribute to the evolution of pancreatitis and lung injury in these models.

1-Cyano-2-hydroxy-3-butene (CHB) has been reported to cause cell death in rat pancreatic acini. In this report, we describe the time-dependent effects of CHB on mouse acinar cell apoptosis and the effects of CHB-induced acinar cell apoptosis on the severity of secretagogue - induced acute pancreatitis in mice. CHB administration to mice resulted in a time-dependent increase in pancreatic apoptosis, which was maximal 12 hours after CHB administration. The severity of pancreatitis was significantly reduced by prior CHB administration and maximal protection was observed when the caerulein injections were started 12 hours after CHB administration. These observations indicate that induction of apoptosis can reduce the severity of pancreatitis and they suggest that induction of pancreatic acinar cell apoptosis may be beneficial in the clinical management of acute pancreatitis.

Two isoforms of the dopamine D2 receptor have been characterized, D2L (long) and D2S (short), generated by alternative splicing from the same gene. They differ by an in-frame insert of 29 amino acids specific to D2L within the putative third intracytoplasmic loop of the receptor. We have previously demonstrated (Montmayeur, J.-P., Guiramand, J., and Borelli, E.(1993) Mol. Endocrinol.7, 161-170) that D2S and D2L, although presenting very similar pharmacological profiles, couple differently to the α-subunit of guanine nucleotide-binding regulatory proteins (G-proteins). In particular, D2L, but not D2S, requires the presence of the α-subunit of the inhibitory G-protein (Gαi2) to elicit greater inhibition of adenylyl cyclase activity. The insert present in D2L must therefore confer the specificity of interaction with Gαi2. Thus, we introduced substitution mutations within the D2L insert. These mutant receptors were expressed in JEG3 cells, a Gαi2-deficient cell line, scoring for those presenting an increased inhibition of adenylyl cyclase by dopamine. Our analysis identified two mutants, S259/262A and D249V, with these properties. These results clearly show that the insert present in D2L plays a critical role in the selectivity for the G-proteins interacting with the receptor.