Trajectories of Lymphocyte Counts in the Early Phase of Acute Pancreatitis Are Associated With Infected Pancreatic Necrosis.

INTRODUCTION: Infected pancreatic necrosis (IPN) is an important complication of acute pancreatitis (AP). Absolute lymphocyte count (ALC) was reported to be associated with immunosuppression and the development of IPN. The aim of this study was to describe the trajectory of ALC during the early phase of AP and assess its association with IPN. METHODS: We retrospectively screened patients with AP admitted to our center between January 2016 and July 2019. The ALC levels for the first 7 days after admission were collected. Group-based trajectory modeling was performed to detect the trajectories. Cox proportional hazards regression model was adopted to identify potential risk factors of IPN. RESULTS: Overall, 292 patients were enrolled for analysis. A triple-group trajectory model was developed, assigning 116 patients to the low-level ALC group, 133 to the medium-level ALC group, and 43 to the high-level ALC group. There was no overall significant difference regarding the incidence of IPN among the 3 groups (P = 0.066). In pairwise comparison, patients in the low-level ALC group had significantly higher incidence of IPN than those in the high-level ALC group (hazard ratio: 3.50; 95% confidence interval: 1.22-10.00, P = 0.020). Length of hospital stay and intensive care unit stay differed significantly among patients with different trajectories (P = 0.042 and 0.033, respectively). DISCUSSION: Despite the fact that the trajectories of ALC is overall insignificant for the development of IPN, patients with persistent low ALC trajectories during the early phase of AP are more likely to develop IPN when compared with patients with high ALC trajectories.

Inhibition of the p38 MAPK pathway attenuates renal injury in pregnant rats with acute necrotizing pancreatitis.

The p38 mitogen-activated protein kinase (MAPK) pathway is an important intracellular signalling pathway that leads to increased expression of pro-inflammatory mediators. Our previous studies have shown that the p38 MAPK pathway was changed in the acute renal injury (ARI) in acute pancreatitis in late pregnancy (APIP), whereas the role of p38 MAPK in APIP-induced ARI has been poorly understood. The present study was undertaken to investigate the participation of the p38 MAPK signalling pathway and the protective effect of SB203580, an inhibitor of p38 MAPK in ARI in APIP. Twenty-four late-gestation SD rats were randomly assigned to four groups: the normal group (N), sham-operated group (SO), acute necrotizing pancreatitis (ANP) group, and p38 MAPK inhibitor (SB203580) treatment group (T). The results showed that serum amylase, lipase, urea, and creatinine levels of p38 inhibitor of T groups were markedly lower than the ANP groups. Additionally, the expression of phosphorylated p38 and myeloperoxidase (MPO), tumour necrosis factor alpha (TNF-α), interleukin (IL)-1ß, IL-6, nuclear factor kappa-B (NF-κB), caspase-3, and terminal deoxynucleotidyl TUNEL-positive cells was markedly lower in the T group than in the ANP group. Our results suggest that SB203580 can inhibit renal injury by inhibiting the P38 MAPK signalling pathway and blocking the inflammatory responses in APIP.

A chimeric recombinant infectious hematopoietic necrosis virus induces protective immune responses against infectious hematopoietic necrosis and infectious pancreatic necrosis in rainbow trout.

Infectious pancreatic necrosis virus (IPNV) and infectious hematopoietic necrosis virus (IHNV) are two common viral pathogens that cause severe economic losses in all salmonid species in culture, but especially in rainbow trout. Although vaccines against both diseases have been commercialized in some countries, no such vaccines are available for them in China. In this study, a recombinant virus was constructed using the IHNV U genogroup Blk94 virus as a backbone vector to express the antigenic gene, VP2, from IPNV via the reverse genetics system. The resulting recombinant virus (rBlk94-VP2) showed stable biological characteristics as confirmed by virus growth kinetic analyses, pathogenicity analyses, indirect immunofluorescence assays and western blotting. Rainbow trout were immunized with rBlk94-VP2 and then challenged with the IPNV ChRtm213 strain and the IHNV Sn1203 strain on day 45 post-vaccination. A significantly higher survival rate against IHNV was obtained in the rBlk94-VP2 group on day 45 post-vaccination (86%) compared with the PBS mock immunized group (2%). Additionally, IPNV loads decreased significantly in the rBlk94-VP2 immunized group in the liver (28.6-fold to 36.5-fold), anterior kidney (21.7-fold to 44.2-fold), and spleen (14.9-fold to 22.7-fold), as compared with the PBS mock control group. The mRNA transcripts for several innate and adaptive immune-related proteins (IFN-γ, IFN-1, Mx-1, CD4, CD8, IgM, and IgT) were also significantly upregulated after rBlk94-VP2 vaccination, and neutralizing antibodies against both IHNV and IPNV were induced on day 45 post-vaccination. Collectively, our results suggest that this recombinant virus could be developed as a vaccine vector to protect rainbow trout against two or more diseases, and our approach lays the foundations for developing live vaccines for rainbow trout.

Mouse model to study pulmonary intravascular macrophage recruitment and lung inflammation in acute necrotizing pancreatitis.

Patients suffering from severe acute pancreatitis (AP) can develop acute lung injury (ALI) with poor outcomes and the mechanisms involved remain incompletely understood. Pulmonary intravascular macrophages (PIMs), which are credited as promoters of ALI, are not constitutively present in humans and rodents; however, there is evidence of PIM recruitment in rodents during some pathological conditions, such as hepatic diseases. Therefore, this study assesses PIM recruitment in the lungs of a mouse model of acute necrotizing pancreatitis (ANP) induced with L-arginine monohydrochloride. Mice were euthanized after 24 h, 72 h and 120 h. Control mice received sham injections of saline. Pancreatic histopathological grading and plasma amylase were used to confirm the development of ANP in L-arginine-treated mice. Histopathological grading of lungs from the ANP mice at 72 h showed increased mononuclear phagocytes in alveolar septa, compared to that from the controls. Lungs from the ANP mice also showed increased numbers of CD68-immunopositive alveolar septal macrophages, suggestive of PIM recruitment, compared to those from the controls. Lungs from the ANP mice showed increased expression of IL-6, IL-10, monocyte chemoattractant protein 1 (MCP-1) and von Willebrand factor compared to those from the controls. The recruitment of CD68-positive septal macrophages was not observed in MCP-1 knockout mice with ANP at 72 h when compared to C57BL/6 wild-type mice. Taken together, we developed a mouse model of PIM recruitment dependent on MCP-1 that allows us to explore their roles in ANP-associated ALI.

The immune imbalance in the second hit of pancreatitis is independent of IL-17A.

BACKGROUND: Severe acute pancreatitis (SAP) is characterised by two distinct clinical phases. Organ dysfunction and death is initially as a result of a systemic inflammatory response syndrome (SIRS). Systemic sepsis from infected pancreatic necrosis characterises the second phase, the so called 'second hit' of acute pancreatitis (AP). An immune imbalance during the second hit is postulated to contribute to the formation of the septic complications that occur in these patients. The pro-inflammatory T-helper (Th) 17 pathway has been shown to be an initiator of early SIRS in AP, however to date its role has not been established in the second hit in AP. METHODS: Thirty-six patients with mild (n = 16), moderate (n = 10) and severe (n = 10) acute pancreatitis were enrolled. Peripheral blood samples were drawn on days 7, 9, 11 and 13 of illness for analysis of routine clinical markers as well as cytokine analysis. Flow cytometry and a IL-17A ELISA was performed to determine cytokine concentrations. RESULTS: There were no significant differences between days 7, 9, 11 and 13 for either the mild/moderate or SAP groups for IL-17A (CBA assay or ELISA), IFN-γ, TNF-α, IL-2 or IL-4. For each of the study days, the mean IL-6 and IL-10 concentrations were significantly higher in the SAP group compared to the mild/moderate group. WCC, CRP and PCT were all significantly higher in severe acute pancreatitis over the study days. CONCLUSIONS: An immune imbalance exists in patients with SAP, however secreted IL-17A is not responsible for the second hit in AP.

Signalling Profiles of Blood Leucocytes in Sepsis and in Acute Pancreatitis in Relation to Disease Severity.

Intracellular signalling in blood leucocytes shows multiple aberrations in acute pancreatitis (AP) complicated by organ dysfunction (OD). We studied whether the aberrations associate with severity of AP and occur in sepsis complicated by OD. The study comprises 14 sepsis patients (11 with shock), 18 AP patients (nine mild; six moderately severe; three severe) and 28 healthy volunteers. Within 48 h after admission to hospital, phosphorylation of nuclear factor-ĸB (NF-ĸB), signal transducers and activators of transcription (STATs) 1,3, and extracellular signal-regulated kinases 1/2 were measured from stimulated or non-stimulated leucocytes using phosphospecific whole blood flow cytometry. In sepsis, as compared with healthy subjects, phosphorylated NF-ĸB levels of monocytes promoted by bacterial lipopolysaccharides, tumour necrosis factor or Escherichia coli cells were lower (P < 0.001 for all), pSTAT1 levels of monocytes promoted by IL-6 were lower (P < 0.05 for all), and STAT3 was constitutively phosphorylated in monocytes, neutrophils and lymphocytes (P < 0.001 for all). In AP, severity was associated with proportions of pSTAT1-positive monocytes and lymphocytes promoted by IL-6 (P < 0.01 for both), constitutive STAT3 phosphorylation in neutrophils (P < 0.05), but not with any of the pNF-ĸB levels. Monocyte pSTAT3 fluorescence intensity, promoted by IL-6, was lower in sepsis and AP patients with OD than in AP patients without OD (P < 0.001). Collectively, signalling aberrations in sepsis with OD mimic those described previously in AP with OD. Possibility that aberrations in STAT1 and STAT3 pathways provide novel markers predicting evolution of OD warrants studies including patients presenting without OD but developing it during follow-up.

Cathepsin B-Mediated Activation of Trypsinogen in Endocytosing Macrophages Increases Severity of Pancreatitis in Mice.

BACKGROUND & AIMS: Acute pancreatitis is characterized by premature intracellular activation of digestive proteases within pancreatic acini and a consecutive systemic inflammatory response. We investigated how these processes interact during severe pancreatitis in mice. METHODS: Pancreatitis was induced in C57Bl/6 wild-type (control), cathepsin B (CTSB)-knockout, and cathepsin L-knockout mice by partial pancreatic duct ligation with supramaximal caerulein injection, or by repetitive supramaximal caerulein injections alone. Immune cells that infiltrated the pancreas were characterized by immunofluorescence detection of Ly6g, CD206, and CD68. Macrophages were isolated from bone marrow and incubated with bovine trypsinogen or isolated acinar cells; the macrophages were then transferred into pancreatitis control or cathepsin-knockout mice. Activities of proteases and nuclear factor (NF)-κB were determined using fluorogenic substrates and trypsin activity was blocked by nafamostat. Cytokine levels were measured using a cytometric bead array. We performed immunohistochemical analyses to detect trypsinogen, CD206, and CD68 in human chronic pancreatitis (n = 13) and acute necrotizing pancreatitis (n = 15) specimens. RESULTS: Macrophages were the predominant immune cell population that migrated into the pancreas during induction of pancreatitis in control mice. CD68-positive macrophages were found to phagocytose acinar cell components, including zymogen-containing vesicles, in pancreata from mice with pancreatitis, as well as human necrotic pancreatic tissues. Trypsinogen became activated in macrophages cultured with purified trypsinogen or co-cultured with pancreatic acini and in pancreata of mice with pancreatitis; trypsinogen activation required macrophage endocytosis and expression and activity of CTSB, and was sensitive to pH. Activation of trypsinogen in macrophages resulted in translocation of NF-kB and production of inflammatory cytokines; mice without trypsinogen activation (CTSB-knockout mice) in macrophages developed less severe pancreatitis compared with control mice. Transfer of macrophage from control mice to CTSB-knockout mice increased the severity of pancreatitis. Inhibition of trypsin activity in macrophages prevented translocation of NF-κB and production of inflammatory cytokines. CONCLUSIONS: Studying pancreatitis in mice, we found activation of digestive proteases to occur not only in acinar cells but also in macrophages that infiltrate pancreatic tissue. Activation of the proteases in macrophage occurs during endocytosis of zymogen-containing vesicles, and depends on pH and CTSB. This process involves macrophage activation via NF-κB-translocation, and contributes to systemic inflammation and severity of pancreatitis.

Association of Systemic Inflammatory and Anti-inflammatory Responses with Adverse Outcomes in Acute Pancreatitis: Preliminary Results of an Ongoing Study.

INTRODUCTION: This paper reports preliminary data of an ongoing study that evaluates the association of systemic inflammatory response (SIRS) with early severe acute pancreatitis (ESAP) and compensatory anti-inflammatory response syndrome (characterized by HLA-DR down-regulation) with infected pancreatic necrosis (IPN). METHODS: Consecutive patients presenting within 72 h of symptom onset with organ dysfunction and/or local complications were included. Following parameters were recorded: demographics, etiology, SIRS, APACHE II, creatinine, BUN. Circulating IL-8, IL-6, IL-10, TNF-alpha concentrations and expression of HLA-DR and IL-10 by qRT-PCR in PBMCs were measured. Strength of associations of cytokine concentration and HLA-DR/IL-10 expression with outcomes was expressed as Hedges' G and relative risk (95% CI). RESULTS: Twenty-eight patients (10 MSAP; 18 SAP) fulfilled inclusion criteria. Twelve patients had ESAP and eight presented with organ failure. Admission SIRS worsened in eight (28.6%) patients over 48 h. Sixteen (57.1%) patients developed primary IPN. Twenty-one (75%) patients had HLA-DR down-regulation during the first week, which persisted to the second week in 12 (42.9%) patients. IL-8, IL-6, and TNF-α progressively increased from healthy controls to MAP to MSAP to SAP. IL-6 and TNF-α was higher in the patients who developed ESAP (p = 0.01 and 0.05, respectively). Patients who died within the first week also had a significantly elevated concentration of IL-6 and TNF-α (p = 0.02 and 0.01, respectively). The relative risk (95% CI) of developing primary IPN with persistent HLA-DR down-regulation till the second week of illness was 11.3 (1.6-82.4; p = 0.01). CONCLUSIONS: Our study objectively demonstrates significant association of ESAP and early mortality with primary cytokine response, and development of IPN with persistent HLA-DR down-regulation.

Early prediction of infected pancreatic necrosis secondary to necrotizing pancreatitis.

To assess the association between the clinical parameters within 48 hours of admission and the occurrence of infected pancreatic necrosis (IPN) during the late phase of necrotizing pancreatitis (NP).All patients were divided into 2 groups, the IPN and non-IPN groups. The clinical data were retrospectively analyzed. Univariate and multivariate logistic regression analyses were performed to evaluate the relationship between clinical parameters and IPN secondary to NP. The performance of each independent variable was plotted by the receiver-operating characteristic (ROC) curve. Consequently, the cut-off level of each independent variable with its sensitivity and specificity was calculated.A total of 215 patients were enrolled in our study. Among them, 87 (40.5%) patients developed IPNs after a median of 13.5 (9.5-23.0) days from admission. Multivariate analysis indicated that the level of hematocrit (HCT) from 40% to 50% (P=.012, odds ratio (OR) = 2.407), HCT over 50% (P < .009, OR = 6.794), blood urea nitrogen (BUN) (P = .040, OR = 1.894), C-reactive protein (CRP) (P = .043, OR = 1.837), and procalcitonin (PCT) (P = .002, OR = 2.559) were independent risk factors of IPN secondary to NP. The ROC cures revealed that the area under the ROC (AUC) of the maximum level of HCT, BUN, CRP, and PCT within 48 hours of admission was 0.687, 0.620, 0.630, and 0.674 respectively. Furthermore, the combination of these 4 individual parameters contributes to a more preferable AUC of 0.789 with a sensitivity of 67.8% and specificity of 77.3%.The maximum levels of PCT, CRP, HCT, and BUN within 48 hours of admission are independent factors of IPN and their combination might accurately predict the occurrence of IPN secondary to NP.

Epigenetic Regulation of Early- and Late-Response Genes in Acute Pancreatitis.

Chromatin remodeling seems to regulate the patterns of proinflammatory genes. Our aim was to provide new insights into the epigenetic mechanisms that control transcriptional activation of early- and late-response genes in initiation and development of severe acute pancreatitis as a model of acute inflammation. Chromatin changes were studied by chromatin immunoprecipitation analysis, nucleosome positioning, and determination of histone modifications in promoters of proinflammatory genes in vivo in the course of taurocholate-induced necrotizing pancreatitis in rats and in vitro in rat pancreatic AR42J acinar cells stimulated with taurocholate or TNF-α. Here we show that the upregulation of early and late inflammatory genes rely on histone acetylation associated with recruitment of histone acetyltransferase CBP. Chromatin remodeling of early genes during the inflammatory response in vivo is characterized by a rapid and transient increase in H3K14ac, H3K27ac, and H4K5ac as well as by recruitment of chromatin-remodeling complex containing BRG-1. Chromatin remodeling in late genes is characterized by a late and marked increase in histone methylation, particularly in H3K4. JNK and p38 MAPK drive the recruitment of transcription factors and the subsequent upregulation of early and late inflammatory genes, which is associated with nuclear translocation of the early gene Egr-1 In conclusion, specific and strictly ordered epigenetic markers such as histone acetylation and methylation, as well as recruitment of BRG-1-containing remodeling complex are associated with the upregulation of both early and late proinflammatory genes in acute pancreatitis. Our findings highlight the importance of epigenetic regulatory mechanisms in the control of the inflammatory cascade.

The immunoproteasomes are key to regulate myokines and MHC class I expression in idiopathic inflammatory myopathies.

Idiopathic inflammatory myopathies (IIMs) are diseases with muscle weakness, morphologically characterized by inflammatory infiltration and increased expression of MHC class I molecule on myofibers. Immunoproteasome, as a proteolytic complex that shapes the repertoire of antigenic peptides, has been previously demonstrated to be over-expressed in IIMs at mRNA level. In this study, we investigated the expression and the function of the immunoproteasome in IIMs in more detail. As shown by immunofluorescence staining, expression of relevant players of the immunoproteasome was detectable in the inflamed skeletal muscle tissue from IIM patients. In fact, two subunits of the immunoproteasome, ß1i or ß5i were upregulated in sporadic inclusion body myositis, immune-mediated necrotizing myopathies and dermatomyositis muscle biopsies and co-localized with the MHC class I expressing myofibers. Double immunofluorescence revealed that both myofibers and muscle infiltrating cells, including CD8+ T-cells and CD68 + macrophages in IIMs expressed ß1i or ß5i. In addition, we have also investigated the role of the immunoproteasome in myoblasts during in vitro inflammatory conditions. Using human primary myoblasts cultures we found that pro-inflammatory cytokines, TNF-α or IFN-γ upregulate ß1i or ß5i. Selective inhibition or depletion of ß5i amplified the TNF-α or IFN-γ mediated expression of cytokines/chemokines (myokines) in myoblasts. Furthermore, we demonstrated that specific inhibitors of ß1i or ß5i reduced the cell surface expression of MHC class I in myoblasts induced by IFN-γ. Taken together, our data suggest that the immunoproteasome is involved in pathologic MHC class I expression and maintenance of myokine production in IIMs. Thus, induction of the immunoproteasome was identified as a pathomechanism underlying inflammation in IIMs.

Resistin-Like Molecule-α Causes Lung Injury in Rats with Acute Pancreatitis by Activating the PI-3K/Akt-NF-κB Pathway and Promoting Inflammatory Cytokine Release.

BACKGROUND: Resistin-like molecule-α (RELMα) has diverse regulatory functions in inflammation, but its role in severe acute pancreatitis (SAP) and acute pancreatitis associated lung injury (APALI) remains unclear. METHODS: SAP was induced in rats. RELMα protein expression was detected in lung tissue of rats to determine the relationship between APALI and RELMα. To investigate the effect of RELMα overexpression or knockdown on APALI, rats were given an intravenous injection of adenovirus vector before SAP induction. Lung and pancreatic samples were harvested 16 h after induction. After detection of RELMα protein levels, the severity of pancreatic and pulmonary injury was scored histologically, and serum and tissue levels of inflammatory mediators were measured. TUNEL assay and immunofluorescence were used to estimate pulmonary apoptosis and endothelial barrier integrity in lung tissue of SAP rats with RELMα knockdown. RESULTS: RELMα expression was significantly up-regulated in APALI and was related to the lung injury index. RELMα overexpression aggravated the release of inflammatory cytokines including interleukin (IL)-1ß, IL-6, IL-8, tumor necrosis factor-α, and serum C-reaction protein; the expression of inflammatory mediators phosphorylated (p)-AKT, p-P65, p-P38 mitogen activated protein kinase, p-extracellular regulated kinase, and intracellular adhesion molecule-1; and lung injury. RELMα knockdown had opposite effects. In addition, RELMα knockdown improved expression of proliferative cellular nuclear antigen, Bcl-2, zonal occluding-1 and Claudin-1 in lung tissue of SAP rats. CONCLUSION: RELMα is associated with lung injury severity in SAP. RELMα augments inflammatory activity by increasing inflammatory cytokine release.

Minocycline inhibits peritoneal macrophages but activates alveolar macrophages in acute pancreatitis.

Minocycline is a tetracycline antibiotic that, in addition to its antimicrobial function, has been reported to possess a relevant anti-inflammatory activity. Its effects have been extensively evaluated in inflammatory-related neurological diseases. Here, we evaluate its effect on the systemic inflammatory response in a model of experimental acute pancreatitis. Minocycline treatment significantly reduced the inflammation in pancreas and mesenterium, had no effect on the adipose tissue inflammation, and increased the inflammatory response in the lung. These differences seem to be related with different effects exerted on peritoneal and alveolar macrophages. In vitro, minocycline reduced the expression of IL-1ß and inhibit the activation of nuclear factor kappa B (NF-κB) on peritoneal macrophages, while it had no effect on alveolar macrophages. Our data indicates that although minocycline may be useful as a tool to control some inflammatory processes, differences on its effects depending on the population of macrophages involved in the process can be expected. In the particular case of acute pancreatitis, it could promote or potentiate inflammation in the lung so that its use does not appear to be recommended.

Efficacy of thymosin α1 and interferon α for the treatment of severe acute pancreatitis in a rat model.

The present study aimed to investigate the effects of treatment with thymosin α1 (TA1) or interferon α (IFNα) following the establishment of severe acute pancreatitis (SAP) in rats. A total of 144 Sprague­Dawley rats were randomly divided into four groups. The rats in all four groups were celiotomized, and the rats in the control group were administered with an intravenous injection of saline. The three other groups were administered with 5% 1 ml/kg sodium taurocholate via the cholangiopancreatic duct. SAP group rats were administered with an intravenous injection of saline; TA1 group rats received 26.7 µg/kg TA1; and interferon α (INFα) group rats received 4.0x105 U/kg IFNα. The rats were anesthetized and blood samples were collected from the animals 3, 12 and 24 h after surgery. The levels of T cell subsets, serum enzyme indicators, cytokines and procalcitonin (PCT) were measured. The general conditions of the rats were observed until sacrifice, and pancreatic and lung tissue samples were sampled for hematoxylin and eosin staining and histological scoring. The expression levels of aspartate transaminase, lactate dehydrogenase, α­amylase (AMY), P­type­amylase, lipase, PCT, tumor­necrosis factor α, interleukin (IL)­4, IL­5, and IL­18 in the TA1 and IFNα­treated rats were significantly lower, compared with those of the SAP rats within the first 24 h of model establishment (P<0.05). The TA1 and IFNα­treated rats exhibited significantly increased levels of CD3+, CD4+ and CD8+ T cells, and an increased ratio of CD4+/CD8+ cells, compared with SAP rats. Histological analysis revealed that the TA1 and IFNα­treated rats exhibited significantly ameliorated pancreas and lung damage, and mortality rates were reduced from 50.0% (6/12) to 25.0% (3/12) and 33.3% (4/12), respectively. The immunomodulatory agents TA1 and IFNα reduced acute inflammation, decreasing cell damage and enhancing immune function and survival rates in the SAP rats.

Pathophysiological mechanisms of acute pancreatitis define inflammatory markers of clinical prognosis.

Development of acute pancreatitis illustrates the need to understand the basic mechanisms of disease progression to drive the exploration of therapeutic options. Cytokines play a major role in the pathogenesis of acute pancreatitis as underlying systemic inflammatory response, tissue damage, and organ dysfunction. However, little is known about circulating concentrations of these inflammatory markers and their real impact on clinical practice. Experimental studies have suggested that the prognosis for acute pancreatitis depends on the degree of pancreatic necrosis and the intensity of multisystem organ failure generated by the systemic inflammatory response. This suggests an intricate balance between localized tissue damage with proinflammatory cytokine production and a systemic anti-inflammatory response that restricts the inappropriate movement of proinflammatory agents into the circulation. Implication of such mediators suggests that interruption or blunting of an inappropriate immune response has the potential to improve outcome. A detailed understanding of pathophysiological processes and immunological aspects in patients with acute pancreatitis is the basis for the development of therapeutic strategies that will provide significant reductions in morbidity and mortality.

Organ-specific monocyte activation in necrotizing pancreatitis in mice.

BACKGROUND: Acute necrotizing pancreatitis (NAP) induces a systemic inflammatory response syndrome. We investigated the underlying changes of monocytes using different activation markers. MATERIALS AND METHODS: A retrograde injection of 2 mL/kg bodyweight of sodium taurocholate into the common bile duct of BALB/c mice induced NAP, whereas sham-operated animals (SOP) were treated with saline. After 6, 12, 24, and 48 h, histologic alterations, pancreatic enzymes, and interleukin 6 in serum, albumin, and myeloperoxidase (MPO) in bronchoalveolar lavage fluid were examined. Isolation of mononuclear cells from the blood, spleen, and liver and the subsequent determination of macrophages (F4/80) and their activation marker CD121b and MHCII (1Ad) were performed by fluorescence-activated cell sorting (FACS analyses). RESULTS: After pancreatitis induction, pancreatic enzymes (amylase: SOP 7008 U/L, NAP 37,044 U/L, P < 0.001) and histologic pancreatic damage (SOP 0.80 ± 1.92, NAP 19.6 ± 0.64, P < 0.001) developed instantly. Pulmonary vascular damage and MPO were detected between 6 and 12 h after onset (6 h albumin SOP 132.0 ± 12.0 µg/mL, NAP 267.2 ± 49.6 µg/mL; P < 0.05; MPO SOP 0.23 ± 0.20 ng/mL, NAP 11.29 ± 3.12 ng/mL, P < 0.01). Blood levels of interleukin 6 increased after 12-24 h (12 h SOP 584 ± 300 pg/mL; NAP 2169 ± 942 pg/mL, P < 0.05), whereas monocytes increased fourfold within 48 h (P < 0.05). Furthermore, pancreatitis increased the percentage of activated monocytes in the blood (6 h and/or 48 h: MHCII (1Ad) 2196%/5.65%; CD121b 51,654%/82,146%). Similar observations were made for monocytes from the liver and spleen. CONCLUSIONS: Although inflammatory mediators increased during 24 h after pancreatitis induction, monocyte activation lasted for at least 48 h. The latter is not limited to blood but also detected in isolated liver and spleen monocytes.

Nicotine ameliorates experimental severe acute pancreatitis via enhancing immunoregulation of CD4+ CD25+ regulatory T cells.

OBJECTIVES: Activation of "nicotinic anti-inflammatory pathway" could reduce severity of inflammation and injury induced by acute pancreatitis. However, the role of regulatory T (Treg) cells in this pathway is unclear. METHODS: Severe acute pancreatitis (SAP) was induced in mice through retrograde injection of 50-µL 2% Na-taurocholate into the pancreatic duct of the mouse. In nicotine treatment group, nicotine (50, 100, and 300 µg/kg) was administered 1 hour before and after SAP operation through intraperitoneal injection. We compared the properties of Treg cell percentage and specific marker such as cytotoxic T-lymphocyte antigen 4 and forkhead box transcription factor forkhead/winged helix transcription factor p3 on Treg using quantitative reverse transcription polymerase chain reaction and flow cytometry. All experiment animal serum cytokines were measured using enzyme-linked immunosorbent assay. One-way analysis of variance was applied to evaluate the experimental data and for statistical comparisons. The survival rate data were analyzed using the log-rank test. RESULTS: Nicotine significantly protected mice from lethal SAP in a dose-dependent fashion by inhibiting tissue injury, digestive enzyme production, and proinflammatory cytokines production. Moreover, nicotine up-regulated the number and suppressive capacity of CD4 CD25 Treg via inducing the expression of immunoregulatory molecules and transforming growth factor ß1 elevation. CONCLUSIONS: Modulating immunoregulation of CD4 CD25 Treg is a critical mechanism for nicotinic anti-inflammatory pathway and it may be feasible to use selective agonists as an immunotherapy for SAP.

Inhibition of Ras signalling reduces neutrophil infiltration and tissue damage in severe acute pancreatitis.

Neutrophil recruitment is known to be a rate-limiting step in mediating tissue injury in severe acute pancreatitis (AP). However, the signalling mechanisms controlling inflammation and organ damage in AP remain elusive. Herein, we examined the role of Ras signalling in AP. Male C57BL/6 mice were treated with a Ras inhibitor (farnesylthiosalicylic acid, FTS) before infusion of taurocholate into the pancreatic duct. Pancreatic and lung tissues as well as blood were collected 24 h after pancreatitis induction. Pretreatment with FTS decreased serum amylase levels by 82% and significantly attenuated acinar cell necrosis, tissue haemorrhage and oedema formation in taurocholate-induced pancreatitis. Inhibition of Ras signalling reduced myeloperoxidase (MPO) levels in the inflamed pancreas by 42%. In addition, administration of FTS decreased pancreatic levels of CXC chemokines as well as circulating levels of interleukin-6 and high-mobility group box 1 in animals exposed to taurocholate. Moreover, treatment with FTS reduced taurocholate-induced MPO levels in the lung. Inhibition of Ras signalling had no effect on neutrophil expression of Mac-1 in mice with pancreatitis. Moreover, FTS had no direct impact on trypsin activation in isolated pancreatic acinar cells. These results indicate that Ras signalling controls CXC chemokine formation, neutrophil recruitment and tissue injury in severe AP. Thus, our findings highlight a new signalling mechanism regulating neutrophil recruitment in the pancreas and suggest that inhibition of Ras signalling might be a useful strategy to attenuate local and systemic inflammation in severe AP.

C-reactive protein may influence decisively the prescription of prophylactic antibiotics in acute pancreatitis: a population-based cohort study.

OBJECTIVES: Prescription of prophylactic antibiotics in acute pancreatitis (AP) is controversial. We aimed to identify the patients' characteristics that may prompt clinicians to prescribe prophylactic antibiotics in AP. METHODS: This single-center retrospective cohort study included 299 consecutive patients with AP from a Portuguese hospital in 2009 to 2010. Logistic regression was used to study the association of patients' characteristics with prescription of prophylactic antibiotics in AP. RESULTS: Persistent organ failure developed in 7% of patients (9/136). The median C-reactive protein at 48 hours after hospital admission was 154 mg/L (interquartile range, 55-271 mg/L). Bedside Index for Severity in AP score greater than or equal to 3 occurred in 14% of patients (42/299). Pancreatic necrosis was diagnosed in 21% of the patients (35/169). Computerized Tomography Severity Index score greater than 3 occurred in 23% of patients (38/169). In-hospital mortality rate was 4% (10/299). Prophylactic antibiotics were prescribed to 14% of patients (42/299). After adjusting for persistent organ failure and Computerized Tomography Severity Index score greater than 3, C-reactive protein at 48 hours after hospital admission greater than or equal to 150 mg/L was significantly associated with higher likelihood of receiving prophylactic antibiotics (odds ratio, 12.2). Prophylactic antibiotics did not improve in-hospital mortality rate (P = 0.637). CONCLUSIONS: C-reactive protein was the most influential in prescribing prophylactic antibiotics in AP. Clinicians may need better tools to support the decision to prescribe prophylactic antibiotics in AP.

Treatment of severe acute pancreatitis and its complications.

Severe acute pancreatitis (SAP), which is the most serious type of this disorder, is associated with high morbidity and mortality. SAP runs a biphasic course. During the first 1-2 wk, a pro-inflammatory response results in systemic inflammatory response syndrome (SIRS). If the SIRS is severe, it can lead to early multisystem organ failure (MOF). After the first 1-2 wk, a transition from a pro-inflammatory response to an anti-inflammatory response occurs; during this transition, the patient is at risk for intestinal flora translocation and the development of secondary infection of the necrotic tissue, which can result in sepsis and late MOF. Many recommendations have been made regarding SAP management and its complications. However, despite the reduction in overall mortality in the last decade, SAP is still associated with high mortality. In the majority of cases, sterile necrosis should be managed conservatively, whereas in infected necrotizing pancreatitis, the infected non-vital solid tissue should be removed to control the sepsis. Intervention should be delayed for as long as possible to allow better demarcation and liquefaction of the necrosis. Currently, the step-up approach (delay, drain, and debride) may be considered as the reference standard intervention for this disorder.