Yue Zeng

Mucosal adherent bacterial dysbiosis in patients with colorectal adenomas

Recent reports have suggested that the gut microbiota is involved in the progression of colorectal cancer (CRC). The composition of gut microbiota in CRC precursors has not been adequately described. To characterize the structure of adherent microbiota in this disease, we conducted pyrosequencing-based analysis of 16S rRNA genes to determine the bacterial profile of normal colons (healthy controls) and colorectal adenomas (CRC precursors). Adenoma mucosal biopsy samples and adjacent normal colonic mucosa from 31 patients with adenomas and 20 healthy volunteers were profiled using the Illumina MiSeq platform. Principal coordinate analysis (PCoA) showed structural segregation between colorectal adenomatous tissue and control tissue. Alpha diversity estimations revealed higher microbiota diversity in samples from patients with adenomas. Taxonomic analysis illustrated that abundance of eight phyla (Firmicutes, Proteobacteria, Bacteroidetes, Actinobacteria, Chloroflexi, Cyanobacteria, Candidate-division TM7, and Tenericutes) was significantly different. In addition, Lactococcus and Pseudomonas were enriched in preneoplastic tissue, whereas Enterococcus, Bacillus, and Solibacillus were reduced. However, both PCoA and cluster tree analyses showed similar microbiota structure between adenomatous and adjacent non-adenoma tissues. These present findings provide preliminary experimental evidence supporting that colorectal preneoplastic lesion may be the most important factor leading to alterations in bacterial community composition.

Transcriptional repression of SOCS3 mediated by IL-6/STAT3 signaling via DNMT1 promotes pancreatic cancer growth and metastasis

BackgroundPrevious studies have investigated the sustained aberrantly activated Interleukin-6 (IL-6)/signal transducer and activator of transcription 3 (STAT3) signaling pathway is crucial for pancreatic cancer growth and metastasis. Suppressor of cytokine signaling 3 (SOCS3), as a key negative feedback regulator of this signaling pathway, is usually down-regulated in various cancers. In the present study, we aim at exploring the biological function and the underlying molecular regulation mechanisms of SOCS3 in pancreatic cancer.MethodsThe expression of SOCS3 and other genes in pancreatic cancer was examined by Quantitative real-time PCR, western blotting and immunohistochemical staining. The interaction between pSTAT3 and DNA Methyltransferase 1 (DNMT1) was investigated by co-immunoprecipitation assay. Luciferase reporter assay was used to investigate the transcriptional regulation of pSTAT3 and DNMT1 on the SOCS3 gene. The effects of SOCS3 on the biological behavior of pancreatic cancer cells were assessed both in vitro and vivo. Furthermore, we performed a comprehensive analysis of the expression of SOCS3 in a pancreatic cancer tissue microarray (TMA) and correlated our findings with pathological parameters and outcomes of the patients.ResultsWe showed that SOCS3 expression was decreased in phosphorylated STAT3 (pSTAT3)-positive tumors and was negatively correlated with pSTAT3 in pancreatic cancer cells. We also found that IL-6/STAT3 promoted SOCS3 promoter hypermethylation by increasing DNMT1 activity; silencing DNMT1 or 5-aza-2-deoxycytidine (5-AZA) treatment could reverse the down-regulation of SOCS3 mediated by IL-6. Using co-immunoprecipitation and luciferase reporter assays, we found that STAT3 recruited DNMT1 to the promoter region of SOCS3 and inhibited its transcriptional activity. Overexpression of SOCS3 significantly inhibited cell proliferation, which may be due to the increase in G1-S phase arrest; overexpression of SOCS3 also inhibited cell migration and invasion as well as tumorigenicity in nude mice. Pancreatic cancer tissue microarray analysis showed that high SOCS3 expression was a good prognostic factor and negatively correlated with tumor volume and metastasis.ConclusionWe demonstrated that activated IL-6/STAT3 signaling could induce SOCS3 methylation via DNMT1, which led to pancreatic cancer growth and metastasis. These data also provided a mechanistic link between sustained aberrantly activated IL-6/STAT3 signaling and SOCS3 down-regulation in pancreatic cancer. Thus, inhibitors of STAT3 or DNMT1 may become novel strategies for treating pancreatic cancer.

Reverse-migrated neutrophils regulated by JAM-C are involved in acute pancreatitis-associated lung injury.

Junctional adhesion molecule-C (JAM-C) plays a key role in the promotion of the reverse transendothelial migration (rTEM) of neutrophils, which contributes to the dissemination of systemic inflammation and to secondary organ damage. During acute pancreatitis (AP), systemic inflammatory responses lead to distant organ damage and typically result in acute lung injury (ALI). Here, we investigated the role of rTEM neutrophils in AP-associated ALI and the molecular mechanisms by which JAM-C regulates neutrophil rTEM in this disorder. In this study, rTEM neutrophils were identified in the peripheral blood both in murine model of AP and human patients with AP, which elevated with increased severity of lung injury. Pancreatic JAM-C was downregulated during murine experimental pancreatitis, whose expression levels were inversely correlated with both increased neutrophil rTEM and severity of lung injury. Knockout of JAM-C resulted in more severe lung injury and systemic inflammation. Significantly greater numbers of rTEM neutrophils were present both in the circulation and pulmonary vascular washout in JAM-C knockout mice with AP. This study demonstrates that during AP, neutrophils that are recruited to the pancreas may migrate back into the circulation and then contribute to ALI. JAM-C downregulation may contribute to AP-associated ALI via promoting neutrophil rTEM.

miR-183 induces cell proliferation, migration, and invasion by regulating PDCD4 expression in the SW1990 pancreatic cancer cell line

The aim of this study was to investigate the function of miR-183 in the SW1990 cancer cell line, and the mechanisms regulating these processes. miRNAs are known to play important roles in cancer cell development. However, the pattern and biological role of miR-183 in pancreatic cancer remain largely unknown. Here, we have reported the reduction in pancreatic cancer cell growth in vitro by miR-183 intervention, by inducing apoptosis and decreasing the Bcl-2 expression. Moreover, miR-183 was observed to enhance pancreatic cancer cell migration and invasion, whereas inhibition of miR-183 caused an opposite effect. miR-183 inhibition was shown to increase E-cadherin expression and decrease N-cadherin expression. These regulatory actions play an important role in the cancer epithelial-mesenchymal transition (EMT). Mechanistically, we demonstrated that the overexpression of miR-183 decreased the expression of PDCD4 (programmed cell death 4) mRNA and protein, and vice versa. This helped to identify PDCD4 as the target genes in pancreatic cancer. In conclusion, our analyses indicated miR-183 to be an important contributor to cell migration. This could also be used as a potential therapeutic target for pancreatic cancer treatment.

Clinical features and treatment of hypertriglyceridemia‐induced acute pancreatitis during pregnancy: A retrospective study

Aim: To analyze the features and treatment of hypertriglyceridemia‐induced acute pancreatitis (HTGP) during pregnancy. Methods: A retrospective study of 21 pregnant women diagnosed with acute pancreatitis (AP) was performed. Patients were divided into acute biliary pancreatitis (ABP), HTGP, and idiopathic groups according to etiology. Results: 95% of the patients were in the third trimester of gestation. The percentage of patients with HTGP was higher than that of ABP (48% vs.14%). The percentage of severe acute pancreatitis (SAP) in the HTGP group was higher than that in the ABP group (40.0% vs.0%). The Ranson scores for moderately severe acute pancreatitis (MSAP) and SAP in the HTGP group were significantly different (2.50 ± 0.58 vs.3.60 ± 0.89, P < 0.05, respectively). The mean serum triglyceride (TG) levels in the MSAP and SAP HTGP groups were not significantly different (18.81 ± 11.13 vs. 30.53 ± 24.20 mmol/L, P > 0.05, respectively). In the HTGP group, there were five patients given plasma exchange therapy and five not. Plasmapheresis decreased the incidence of systemic inflammatory response syndrome (SIRS) from 100% to 28.6% and the TG level from 20.36 ± 7.41 mmol/L to 5.23 ± 3.62 mmol/L (P < 0.05). The length of hospitalization of the plasmapheresis group was shorter than that of the nonplasmapheresis group (17.3 ± 6.7 days vs. 37.0 ± 20.8 days). Conclusions: Plasma exchange may be safe and effectively administered for HTGP patients during pregnancy with SIRS or multiple organ dysfunction syndrome (MODS). J. Clin. Apheresis 31:571–578, 2016.

Impact of Hypertriglyceridemia on the Outcome of Acute Biliary Pancreatitis

Background:Hypertriglyceridemia (HTG) is a well-recognized cause of acute pancreatitis (AP). However, the role of HTG in modulating disease course remains to be cleared. We aimed to explore the impact of HTG on the outcome of acute biliary pancreatitis (ABP). Methods:A total of 90 ABP patients with HTG were enrolled in this study and were divided into 3 groups based on ABP severity: mild AP, moderately severe AP (MSAP) and severe AP (SAP), according to the modified Atlanta classification. Besides, patients were divided into type I, II and III HTG groups based on the triglyceride (TG) level, according to the national cholesterol education program (NCEP). Disease severity, Ranson score and complications were recorded and analyzed. Logistic regression analysis was performed to screen predictive risk factors of severe outcomes. Results:There were 23 patients with SAP, 41 with MSAP and 26 with mild AP among the ABP patients with HTG. Significant differences were observed in the obesity, plasma TG level and Ranson score among groups. The risk of developing MSAP in ABP patients with type II and III HTG was increased. Similarly, the risk of developing SAP in ABP patients with type III HTG was also enhanced. The incidence of systemic, especially respiratory failure, and local complications in type III HTG group were significantly higher than normal lipid group. Conclusions:High TG level (≥2.26 mmol/L or 200.11 mg/mL) may be a risk factor for severe ABP and the development of systemic and local complications in ABP.

Dysbiosis of Intestinal Microbiota and Decreased Antimicrobial Peptide Level in Paneth Cells during Hypertriglyceridemia-Related Acute Necrotizing Pancreatitis in Rats.

Hypertriglyceridemia (HTG) aggravates the course of acute pancreatitis (AP). Intestinal barrier dysfunction is implicated in the pathogenesis of AP during which dysbiosis of intestinal microbiota contributes to the dysfunction in intestinal barrier. However, few studies focus on the changes in intestine during HTG-related acute necrotizing pancreatitis (ANP). Here, we investigated the changes in intestinal microbiota and Paneth cell antimicrobial peptides (AMPs) in HTG-related ANP (HANP) in rats. Rats fed a high-fat diet to induce HTG and ANP was induced by retrograde injection of 3.5% sodium taurocholate into biliopancreatic duct. Rats were sacrificed at 24 and 48 h, respectively. Pancreatic and ileal injuries were evaluated by histological scores. Intestinal barrier function was assessed by plasma diamine oxidase activity and D-lactate level. Systemic and intestinal inflammation was evaluated by tumor necrosis factor alpha (TNFα), interleukin (IL)-1β, and IL-17A expression. 16S rRNA high throughput sequencing was used to investigate changes in intestinal microbiota diversity and structure. AMPs (α-defensin5 and lysozyme) expression was measured by real-time polymerase chain reaction (PCR) and immunofluorescence. The results showed that compared with those of normal-lipid ANP (NANP) groups, the HANP groups had more severe histopathological injuries in pancreas and distal ileum, aggravated intestinal barrier dysfunction and increased TNFα, IL-1β, and IL-17A expression in plasma and distal ileum. Principal component analysis showed structural segregation between the HANP and NANP group. α-Diversity estimators in the HANP group revealed decreased microbiota diversity compared with that in NANP group. Taxonomic analysis showed dysbiosis of intestinal microbiota structure. In the HANP group, at phyla level, Candidatus_Saccharibacteria and Tenericutes decreased significantly, whereas Actinobacteria increased. At genus level, Allobaculum, Bifidobacterium, and Parasutterella increased significantly, while Alloprevotella, Anaerotruncus, Candidatus_Saccharimonas, Christensenellaceae_R-7_group, Rikenellaceae_RC9_gut_group, Ruminiclostridium_5, Ruminococcaceae_UCG-005, and Ruminococcaceae_UCG-014 decreased. Compared with those in the NANP rats, mRNA expression of lysozyme and α-defensin5 and protein expression of lysozyme decreased significantly in the HANP rats. Moreover, in the NANP rats and the HANP rats, Allobaculum abundance was inversely correlated with lysozyme expression, while Anaerotruncus abundance was positively correlated with it by Spearman test. In conclusion, intestinal microbiota dysbiosis and decreased AMPs of Paneth cells might participate in the pathogenesis of intestinal barrier dysfunction in HANP.

Hyperlipidemic versus normal-lipid acute necrotic pancreatitis: proteomic analysis using an animal model.

Objectives Hyperlipidemia is associated with a variety of pancreatic diseases. However, the underlying pathophysiology and molecular mechanisms between hyperlipidemia and acute pancreatitis remain undefined. Gel electrophoresis and mass spectrometry can be used in proteomic analysis to elucidate these mechanisms. Methods A comparative proteomic analysis was conducted to identify proteins that were altered in pancreases of hyperlipidemic acute necrotic pancreatitis rats compared with those of normal-lipid acute necrotic pancreatitis rats. A comparative proteomic approach using a hyperlipidemic rat model was used. Results Thirty-nine differentially expressed proteins were significantly changed in pancreatic samples from hyperlipidemic acute necrotic pancreatitis rats. Differentially expressed proteins in hyperlipidemic pancreatitis include pancreatic proteolytic enzymes, such as lipase, amylase, carboxypolypeptidase, and &agr;-1-antiproteinase; endoplasmic reticulum stress–related proteins; and calcium influx–related proteins including protein disulfide isomerase, calreticulin, annexin A, glucose-regulated protein 78, heat shock protein 60, and peroxiredoxin. Other proteins associated with DNA replication and damage repair, apoptosis, cell metabolism, circulatory dysfunction, and signal transduction were identified in hyperlipidemic pancreatitis. Conclusions Hyperlipidemia intensifies acute necrotic pancreatitis through various ways. These enzymes may be putative biomarkers of hyperlipidemic acute necrotic pancreatitis.

Hyperlipidemic versus healthy pancreases: A proteomic analysis using an animal model

Hyperlipidemia is associated with a variety of pancreatic diseases; however, the underlying pathophysiology and molecular mechanisms remain undefined. Here, we performed a comparative proteomic analysis of pancreatic tissue obtained from hyperlipidemic rats to identify proteins that may be involved in mediating hyperlipidemia‐associated pancreatic injury. Rats were fed a high‐fat diet to induce hyperlipidemia. Control rats were fed a diet with normal fat content. Pancreatic tissue samples were obtained after 6 or 12 weeks and comparative proteomic analysis, using gel electrophoresis and mass spectrometry, was conducted to identify proteins, the expression of which were altered in pancreases from hyperlipidemic compared with control rat pancreases. The expression levels of 3 of 13 proteins were significantly altered in pancreatic samples from hyperlipidemic rats. Alpha‐amylase and arginase II were dysregulated by more than twofold. These modulations persisted in pancreatic tissue obtained from late‐stage hyperlipidemic rats. The levels of alpha‐amylase and arginase II were significantly altered in pancreases obtained from rats with hyperlipidemia. These enzymes may be putative biomarkers of hyperlipidemia‐mediated pancreatic injury.

Dysbiosis of intestinal microbiota and decrease in paneth cell antimicrobial peptide level during acute necrotizing pancreatitis in rats

Objectives Intestinal barrier dysfunction plays an important role in acute necrotizing pancreatitis (ANP) and intestinal microbiota dysbiosis was involved in intestinal barrier failure. Paneth cells protect intestinal barrier and are associated with intestinal microbiota. Here, we investigated changes in intestinal microbiota and antimicrobial peptides of Paneth cells in ileum during ANP. Methods Rats with ANP were established by retrograde injection of 3.5% sodium taurocholate into biliopancreatic duct and sacrificed at 24h and 48h, respectively. Injuries of pancreas and distal ileum were evaluated by histopathological score. Intestinal barrier function was assessed by plasma diamine oxidase activity (DAO) and D-lactate. Systemic and intestinal inflammation was evaluated by TNFα, IL-1β and IL-17A concentration by ELISA, respectively. 16S rRNA high throughput sequencing on fecal samples was used to investigate the changes in intestinal microbiota in the ANP group at 48h. Lysozyme and α-defensin5 were measured by real-time PCR, western blot and immunofluoresence. Results ANP rats had more severe histopathological injuries in pancreas and distal ileum, injured intestinal barrier and increased expression of TNFα, IL-1β and IL-17A in plasma and distal ileum compared with those of the sham-operated (SO) group. Principal component analysis (PCA) showed structural segregation between the SO and ANP groups. Operational taxonomic unit (OTU) number and ACE index revealed decreased microbiota diversity in the ANP group. Taxonomic analysis showed dysbiosis of intestinal microbiota structure. At phyla level, Saccharibacteria and Tenericutes decreased significantly. At genus level, Escherichia-Shigella and Phascolarctobacterium increased significantly, while Candidatus_Saccharimonas, Prevotellaceae_UCG-001, Lachnospiraceae_UCG-001, Ruminiclostridium_5 and Ruminococcaceae_UCG-008 decreased significantly. Lysozyme and α-defensin5 mRNA expression levels decreased significantly in ANP group at 48h. Protein expression of lysozyme decreased in ANP groups at 24h and 48h. Meanwhile, the relative abundance of Escherichia-Shigella correlated inversely with the decrease in lysozyme. Conclusion The disorder in intestinal microbiota and decreases of Paneth cell antimicrobial peptides might participate in the pathogenesis of intestinal barrier dysfunction during ANP.