MicroRNA-130b Suppresses Malignant Behaviours and Inhibits the Activation of the PI3K/Akt Signaling Pathway by Targeting MET in Pancreatic Cancer.

There has been interested in the microRNAs' roles in pancreatic cancer (PC) cell biology, particularly in regulating pathways related to tumorigenesis. The study aimed to explore the hub miRNAs in PC and underlying mechanisms by bioinformatics and fundamental experiments. RNA datasets collected from the Gene Expression Omnibus were analysed to find out differentially expressed RNAs (DERNAs). The miRNA-mRNA and protein-protein interaction (PPI) networks were built. The clinicopathological features and expressions of hub miRNAs and hub mRNAs were explored. Dual-luciferase reporter gene assay was performed to assess the interaction between microRNA and target gene. RT-qPCR and western blot were employed to explore RNA expression. The roles of RNA were detected by CCK-8 test, wound healing, transwell, and flow cytometry experiment. We verified 40 DEmiRNAs and 1613 DEmRNAs, then detected a total of 69 final functional mRNAs (FmRNAs) and 23 DEmiRNAs. In the miRNA-mRNA networks, microRNA-130b (miR-130b) was the hub RNA with highest degrees. Clinical analysis revealed that miR-130b was considerably lower expressed in cancerous tissues than in healthy ones, and patients with higher-expressed miR-130b had a better prognosis. Mechanically, miR-130b directly targeted MET in PC cells. Cell functional experiments verified that miR-130b suppressed cell proliferation, migration, promoted apoptosis, and inhibited the PI3K/Akt pathway by targeting MET in PC cells. Our findings illustrated the specific molecular mechanism of miR-130b regulating PC progress. The miR-130b/MET axis may be an alternative target in the therapeutic intervention of PC and provide an opportunity to deepen our understanding of the pathogenesis of PC.

Exosomal miR-19a decreases insulin production by targeting Neurod1 in pancreatic cancer associated diabetes.

BACKGROUND: New onset diabetes mellitus demonstrates a roughly correlation with pancreatic cancer (PaC), which is unique in PaC and was named as PaC-induced DM, but the inner mechanism remains unclear. Exosomes mediate intercellular communication and bearing microRNAs might be direct constituent of effect in target cells. METHODS AND RESULTS: The isolated exosomes from PaC cells were used to treat pancreatic ß cells or the primary mice islets, and the glucose stimulated insulin secretions were measured. We validated the exosomal miR-19a from PaC cells to be an important mediator in the down regulation of insulin secretion by targeting Neurod1, the validated gene involved in insulin secretion, by using the quantitative real-time PCR, western blot, and promoter luciferase activity. The relative insulin, cAMP and Ca2+ expressions were also assayed to verify the inverse correlation between cancerous miR-19a and pancreatic islets Neurod1. CONCLUSIONS: Our study indicated that signal changes between cancer cells and ß cells via exosomes might be important in the pathogenesis of PaC-induced DM and supplemented the pathogenesis of PaC-induced DM and provide a possible access of PaC screening strategy.

MicroRNA-221-3p is related to survival and promotes tumour progression in pancreatic cancer: a comprehensive study on functions and clinicopathological value.

BACKGROUND: The microRNA miR-221-3p has previously been found to be an underlying biomarker of pancreatic cancer. However, the mechanisms of miR-221-3p underlying its role in pancreatic cancer pathogenesis, proliferation capability, invasion ability, drug resistance and apoptosis and the clinicopathological value of miR-221-3p have not been thoroughly studied. METHODS: Based on microarray and miRNA-sequencing data extracted from Gene Expression Omnibus (GEO), The Cancer Genome Atlas (TCGA), relevant literature, and real-time quantitative PCR (RT-qPCR), we explored clinicopathological features and the expression of miR-221-3p to determine its clinical effect in pancreatic cancer. Proliferation, migration, invasion, apoptosis and in vitro cytotoxicity tests were selected to examine the roles of mir-221-3p. In addition, several miR-221-3p functional analyses were conducted, including Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) and Protein-protein interaction (PPI) network analyses, to examine gene interactions with miR-221-3p. RESULTS: The findings of integrated multi-analysis revealed higher miR-221-3p expression in pancreatic cancer tissues and blood than that in para-carcinoma samples (SMD of miR-221-3p: 1.52; 95% CI 0.96, 2.08). MiR-221-3p is related to survival both in pancreatic cancer and pancreatic ductal adenocarcinoma patients. Cell experiments demonstrated that miR-221-3p promotes pancreatic cancer cell proliferation capability, migration ability, invasion ability, and drug resistance but inhibits apoptosis. Further pancreatic cancer bioinformatics analyses projected 30 genes as the underlying targets of miR-221-3p. The genes were significantly distributed in diverse critical pathways, including microRNAs in cancer, viral carcinogenesis, and the PI3K-Akt signalling pathway. Additionally, PPI indicated four hub genes with threshold values of 5: KIT, CDKN1B, RUNX2, and BCL2L11. Moreover, cell studies showed that miR-221-3p can inhibit these four hub genes expression in pancreatic cancer. CONCLUSIONS: Our research revealed that pancreatic cancer expresses a high-level of miR-221-3p, indicating a potential miR-221-3p role as a prognosis predictor in pancreatic cancer. Moreover, miR-221-3p promotes proliferation capacity, migration ability, invasion ability, and drug resistance but inhibits apoptosis in pancreatic cancer. The function of miR-221-3p in the development of pancreatic cancer may be mediated by the inhibition of hub genes expression. All these results might provide an opportunity to extend the understanding of pancreatic cancer pathogenesis.

Comparison of esophageal motility in gastroesophageal reflux disease with and without globus sensation.

BACKGROUNDS AND AIMS: This study aimed to compare the esophageal motility between gastroesophageal reflux disease (GERD) patients with typical symptoms but without globus sensation and GERD patients only with globus symptoms. METHODS: A total of 57 consecutive GERD patients diagnosed by endoscopy or by 24-hour pH monitoring between May 2013 and September 2015 were included retrospectively into the study. The patients were grouped based on the presence or absence of globus. Thirty patients presented with typical reflux symptoms but without globus were assigned to the typical GERD group and 27 patients only with globus symptom were assigned to the globus GERD group. All patients underwent esophageal high resolution manometry (HRM) and the differences in esophageal motility between the two groups were analyzed. RESULTS: The globus GERD group showed a significantly greater lower esophageal sphincter (LES) length, LES basal pressure and upper esophageal sphincter (UES) residual pressure than that of the typical GERD group (3.47 ± 0.76 vs. 2.65 ± 0.62 cm, 21.71 ± 9.68 vs. 16.04 ± 8.49 mmHg, 7.30 ± 4.42 vs. 4.12 ± 2.92 mmHg, all p < 0.05). There was no significant difference between the two groups in terms of the distal wave amplitude, mean wave duration, distal contractile integral (DCI), contractile front velocity (CFV), distal latency (DL), integrated relaxation pressure (IRP) and UES basal pressure. The incidence of esophageal dysmotility in the globus GERD group (33.3%) was higher than in the typical GERD group (23.3%). There was no significant difference in esophageal acid exposure of the non-erosive gastroesophageal reflux disease (NERD) patients between the two groups. CONCLUSIONS: Globus GERD patients have a higher UES residual pressure, longer LES length, higher LES basal pressure and greater esophageal dysmotility than typical GERD patients. HRM is useful in evaluating esophageal motility of GERD patients.

Identification of Circulating MiR-25 as a Potential Biomarker for Pancreatic Cancer Diagnosis.

BACKGROUND: The aim of this study was to identify novel microRNAs for potential use in the diagnosis of pancreatic cancer (PaC). METHODS: A total of 1063 serum samples from 303 patients with PaC were collected, and the expression level of miR-25 was measured using quantitative real-time PCR (qRT-PCR). RESULTS: We found that miR-25 had significant diagnostic value for the differential diagnosis of PaC in normal controls with an AUC (the area under the ROC curve) of 0.915 (95% CI: 0.893-0.937) that was significantly higher compared with an AUC of 0.725 for serum tumor marker carcinoembryonic antigen (CEA) and an AUC of 0.844 for CA19-9. CONCLUSIONS: These data suggest that serum miR-25 has strong potential as a novel biomarker for the early detection of PaC.

Pancreatic cancer-secreted miR-155 implicates in the conversion from normal fibroblasts to cancer-associated fibroblasts.

Cancer-associated fibroblasts (CAF) are a major constituent of the pancreatic cancer microenvironment and that the meaning is as intended. Pancreatic cancer cells can induce normal fibroblasts to convert into CAF and, reciprocally, CAF promote tumor invasions and proliferations. The mechanism of the conversion from normal fibroblasts (NF) to CAF remains unclear. MicroRNA are short non-coding RNA involved in the post-transcription gene regulation, which have been defined as an imperative controller in tumor invasions, proliferations and colony formations. Microvesicles (MV) have been proved to be an important mediator of intercellular communication and can selectively transport secreted microRNA from a donor cell into a recipient cell. In this study, we isolated primary pancreatic fibroblasts from wild type C57 mice and co-cultured them with pancreatic cancer cell lines, BxPC-3 and SW1990, and observed the conversion from NF to CAF, or at least CAF-like cells. This phenomenon could also be replicated in primary fibroblasts treated with MV separated from a cancer cell media. We identified that miR-155 was upregulated in PaC-derived MV and we confirmed that normal fibroblasts could convert into CAF after MV containing miR-155 had been taken up. TP53INP1 is a target of miR-155 in fibroblasts and a downregulation of TP53INP1 protein levels could contribute to the fibroblasts' activation. These results indicated that pancreatic cancer cells might reprogram normal adjacent fibroblasts into CAF by means of secreted MV containing miR-155. Targeting the circulating microRNA might be a potential therapy for malignant tumors.

Role of the potassium chloride cotransporter isoform 2-mediated spinal chloride homeostasis in a rat model of visceral hypersensitivity.

Visceral hypersensitivity represents an important hallmark in the pathophysiology of irritable bowel syndrome (IBS), of which the mechanisms remain elusive. The present study was designed to examine whether cation-chloride cotransporter (CCC)-mediated chloride (Cl(-)) homeostasis of the spinal cord is involved in chronic stress-induced visceral hypersensitivity. Chronic visceral hypersensitivity was induced by exposing male Wistar rats to water avoidance stress (WAS). RT-PCR, Western blotting, and immunohistochemistry were used to assess the expression of CCCs in the spinal cord. Patch-clamp recordings were performed on adult spinal cord slices to evaluate Cl(-) homeostasis and Cl(-) extrusion capacity of lamina I neurons. Visceral sensitivity was estimated by measuring the abdominal withdrawal reflex in response to colorectal distension (CRD). After 10 days of WAS exposure, levels of both total protein and the oligomeric form of the K(+)-Cl(-) cotransporter isoform 2 (KCC2), but not Na(+)-K(+)-2Cl(-) transporter isoform 1 (NKCC1), were significantly decreased in the dorsal horn of the lumbosacral spinal cord. The downregulation of KCC2 resulted in a depolarizing shifted equilibrium potential of GABAergic inhibitory postsynaptic current and impaired Cl(-) extrusion capacity in lamina I neurons of the lumbosacral spinal cord from WAS rats. Acute noxious CRD disrupted spinal KCC2 expression and function 2 h after the final distention in sham rats, but not in WAS rats. Pharmacological blockade of KCC2 activity by intrathecal injection of a KCC2 inhibitor [(dihydroindenyl)oxy] alkanoic acid enhanced visceral nociceptive sensitivity in sham rats, but not in WAS rats. These results suggest that KCC2 downregulation-mediated impairment of spinal cord Cl(-) homeostasis may play an important role in chronic stress-induced visceral hypersensitivity.

Exploring the Wnt pathway-associated LncRNAs and genes involved in pancreatic carcinogenesis driven by Tp53 mutation.

PURPOSE: Study the contribution of long non-coding RNAs (lncRNAs) to progression of pancreatic intraepithelial neoplasia (PanIN) to pancreatic ductal adenocarcinoma (PDAC). METHODS: We explored lncRNAs profilings in PanIN cell line (SH-PAN) isolated from Pdx-1-Cre; LSL-Kras (G12D/+) mice and PDAC cell line (DT-PCa) isolated from Pdx-1-Cre; LSL- Kras (G12D/+) ; LSL- Tp53 (R172H/+) mice by lncRNAs microarray, and detected expression of lncRNAs and genes in PDAC by Real-time PCR, Western blot, ChIP and immunohistochemistry. RESULTS: Eight lncRNAs and five protein-coding genes, associated with Wnt pathway, were identified with more than five-fold changes between DT-PCa cells and SH-PAN cells. Of them, lincRNA1611 and Ppp3ca were validated significantly high expression in DT-PCa cells and in 22 of 26 fresh resected human PDAC tissues, compared to SH-PAN cells and normal pancreatic tissues, respectively. Moreover, Tp53 mutation status displayed a positive correlation with lincRNA1611 or Ppp3ca level. Immunohistochemical staining for Ppp3ca was weak or lack in 91 of 107 normal pancreatic tissues, 24 of 29 PanIN-I and 13 of 16 PanIN-II tissues, however, was strong in 10 of 27 PanIN-III and 62 of 107 PDAC tissues post operation. CONCLUSIONS: LincRNA1611 and Ppp3ca were high expression in PDAC and may serve as new potential targets for intervention of the disease.

An all-in-one therapeutic platform for the treatment of resistant Helicobacter pylori infection.

Helicobacter pylori (H. pylori) infection is a major cause of gastric diseases. Currently, bismuth-based quadruple therapy is widely adopted for eradicating H. pylori infection. However, this first-line strategy faces several challenges such as drug resistance, intestinal dysbacteriosis, and patients' poor compliance. To overcome these problems, an all-in-one therapeutic platform (CLA-Bi-ZnO2@Lipo) that composed of liposomes loading clarithromycin (CLA), Bi, and ZnO2 hybrid nanoparticles was developed for eradicating multidrug-resistant (MDR) H. pylori. The in vitro and in vivo results showed that CLA-Bi-ZnO2@Lipo could target the infection-induced inflammatory mucosa through liposome mediated nanoparticle-tissue surface charge interaction and quickly respond to the gastric acid environment to release CLA, Bi3+, Zn2+, and H2O2. By oral administration per day, the acid triggered decomposition of CLA-Bi-ZnO2@Lipo could significantly increase intragastric pH to 6 within 30 min; The released CLA, Zn2+, and H2O2 further exerted synergistical anti-bacterial effects in which a ∼2 order higher efficacy in reducing MDR H. pylori burden was achieved in comparison with standard quadruple therapy (p < 0.05); The released Zn2+ and Bi3+ could also alleviate mucosal inflammation. Most importantly, the CLA-Bi-ZnO2@Lipo exhibited superior biosafety and nearly no side effects on intestinal flora. Overall, this study developed a highly integrated and safe anti-MDR H. pylori agent which had great potential to be used as an alternative treatment for MDR H. pylori eradication.

Contribution of a Circulating 2'-O-methylated MicroRNA Panel to the Diagnosis of Pancreatic Ductal Adenocarcinoma.

Background: We conducted an assessment of 2'-O-methylated (2'OMe) microRNAs (miRNAs) present in the circulation of individuals suffering from pancreatic ductal adenocarcinoma (PDAC). Subsequently, we devised a set of circulating 2'OMe miRNAs that can be utilized for the screening of PDAC patients within a group at increased risk. Methods: A four-step, multicenter research was carried out. The initial screening phase involved analyzing 10 samples from patients with pancreatic ductal adenocarcinoma (PDAC) and 10 specimens from donors who were in good health. RNA sequencing was performed on these specimens after pre-treatment via NaIO4. The instruction and confirmation phases involved the use of 2'OMe miRNA profiling and multivariate analysis to examine applicant 2'OMe miRNAs in a sample of 248 individuals. In a prospective registration population of 135 individuals, a clinical screening panel was created and confirmed. The performance of individual 2'OMe miRNAs or the biomarker panel was evaluated using the receiver operating characteristic curve. Results: Abnormal circulating 2'OMe miRNAs were detected in individuals suspected of pancreatic ductal adenocarcinoma (PDAC). A circulating panel of 3-2'OMe miRNAs, namely miR-28-3p, miR-143-3p, and miR-151a-3p, was subsequently identified. These miRNAs continually exhibited up-regulation in plasma samples of patients with pancreatic ductal adenocarcinoma (PDAC). The panel's area under the curve (AUC) was 1.0 in the experimental group and 0.928 in the verification cohort when comparing PDAC patients in all clinical stages to normal controls. During the application study, both the specificity and sensitivity were found to be 75.00% and 89.72% respectively, with an AUC value of 0.868. In the comparison between early-stage (I-II) PDAC patients and control subjects, the panel demonstrated an AUC of 1.0 in the training cohort and 0.924 in the validation population. In the application group the AUC was 0.810 (95% CI 0.729-0.876) in comparison to the high-risk symptomatic group. Conclusion: Abnormal circulating 2'OMe miRNAs were detected in individuals with pancreatic ductal adenocarcinoma (PDAC). Consequently, we devised a 2'OMe miRNA biological marker panel that holds promise as an initial detection tool for PDAC.

Role of voltage gated Ca2+ channels in rat visceral hypersensitivity change induced by 2,4,6-trinitrobenzene sulfonic acid.

BACKGROUND: Visceral pain is common symptom involved in many gastrointestinal disorders such as inflammatory bowel disease. The underlying molecular mechanisms remain elusive. We investigated the molecular mechanisms and the role for voltage gated calcium channel (VGCC) in the pathogenesis in a rat model of 2,4,6-trinitrobenzenesulfonic acid (TNBS) induced visceral inflammatory hypersensitivity. RESULTS: Using Agilent cDNA arrays, we found 172 genes changed significantly in dorsal root ganglia (DRG) of TNBS treated rats. Among these changed genes, Cav1.2 and Cav2.3 were significantly up-regulated. Then the RT-PCR and Western blot further confirmed the up-regulation of Cav1.2 and Cav2.3. The whole cell patch clamp recording of acutely dissociated colonic specific DRG neurons showed that the peak IBa density was significantly increased in colonic neurons of TNBS treated rats compared with control rats (-127.82 ± 20.82 pA/pF Vs -91.67 ± 19.02 pA/pF, n = 9, *P < 0.05). To distinguish the different type of calcium currents with the corresponding selective channel blockers, we found that L-type (-38.56 ± 3.97 pA/pF Vs -25.75 ± 3.35 pA/pF, n = 9, * P < 0.05) and R-type (-13.31 ± 1.36 pA/pF Vs -8.60 ± 1.25 pA/pF, n = 9, * P < 0.05) calcium current density were significantly increased in colonic DRG neurons of TNBS treated rats compared with control rats. In addition, pharmacological blockade with L-type antagonist (nimodipine) and R-type antagonist (SNX-482) with intrathecal injection attenuates visceral pain in TNBS induced inflammatory visceral hypersensitivity. CONCLUSION: Cav1.2 and Cav2.3 in colonic primary sensory neurons play an important role in visceral inflammatory hyperalgesia, which maybe the potential therapeutic targets.

Upregulation of transgelin is an independent factor predictive of poor prognosis in patients with advanced pancreatic cancer.

Transgelin is a known actin-binding protein, which plays a role in regulating the functions of smooth muscle cells or fibroblasts. Recent evidence indicates that transgelin is involved in diverse human cancers, yet its role in pancreatic cancer remains unclear. We therefore evaluated the expression characteristics and function of transgelin in pancreatic cancer. Immunohistochemical analysis of benign (n = 30 patients) and malignant (n = 114 patients) pancreatic ductal cells showed significantly higher transgelin staining in malignant cells. Lymph node metastasis (P = 0.026) and diabetes (P = 0.041) were shown to significantly correlate with transgelin protein expression. Patients with high transgelin expression showed a shorter 5-year overall survival and a lower tumor-specific survival than those with low transgelin expression. Multivariate analysis revealed that transgelin was an independent factor affecting pancreatic tumor-specific survival (P = 0.025). In vitro, RNA interference-mediated transgelin knockdown resulted in inhibition of pancreatic cancer cell proliferation, migration and invasion. Depletion of transgelin expression could suppress pancreatic tumorigenicity and tumor growth in vivo, and produce enhanced cytotoxic effects of gemcitabine on pancreatic cancer cells both in vitro and in vivo. Our results indicate that transgelin plays a promoting role in tumor progression, and appears to be a novel prognostic marker for advanced pancreatic cancer.

Associations of Gastrointestinal Tract Tumor Necrosis Factor Receptor-Associated Factor 6 Expression with Clinical Features and Prognosis of Eosinophilic Gastroenteritis.

BACKGROUND: Few studies have been conducted to explore the expression of tumor necrosis factor receptor-associated factor 6 in eosinophilic gastroenteritis patients. Therefore, the expression profile of tumor necrosis factor receptor-associated factor 6 in the gastrointestinal tract of eosinophilic gastroenteritis patients and its associations with clinical features were explored in this study. METHODS: Thirty-four eosinophilic gastroenteritis patients who presented in Ruijin Hospital from December 2012 to May 2019 and had accepted gastrointestinal endoscopic examinations were recruited. Medical records and endoscopic biopsies were collected, and the prognosis was followed up by telephone. Healthy persons were selected as the control group. Hematoxylin and eosin and immunohistochemical staining were performed in both eosinophilic gastroenteritis patients and healthy persons. The final results were analyzed by skilled pathologists, and mean optical density values of tumor necrosis factor receptor-associated factor 6 were calculated by Image J software. Final results were analyzed by Statistical Package for the Social Sciences software 22.0. RESULTS: Thirty-four patients (mean age: 25.56 ± 21.14 years, 61.76% males) were recruited for this study. There was no significant difference in tumor necrosis factor receptor-associated factor 6 mean optical density values of gastric tissues in eosinophilic gastroenteritis patients and healthy people (0.22 ± 0.16 vs. 0.14 ± 0.05, P > .05). Eosinophilic gastroenteritis patients had a significantly lower level of intestinal tumor necrosis factor receptor-associated factor 6 mean optical density values than that of healthy people (0.16 ± 0.05 vs. 0.23 ± 0.06, P < .05). Intestinal tumor necrosis factor receptor-associated factor 6 mean optical density values negatively linearly correlated with serum interleukin-10 level (r = -0.618, P = .043 < .05). There were no differences between eosinophilic gastroenteritis patients with or without relapse regarding the expression level of intestinal tumor necrosis factor receptor-associated factor 6 (P = .227 > .05). CONCLUSION: Patients with eosinophilic gastroenteritis might have a deficiency of intestinal tumor necrosis factor receptor-associated factor 6 compared to healthy controls.

Serum microRNA expression profile as a biomarker in the diagnosis and prognosis of pancreatic cancer.

BACKGROUND: Detection of pancreatic cancer (PaC), particularly at early stages, remains a great challenge owing to lack of specific biomarkers. We sought to identify a PaC-specific serum microRNA (miRNA) expression profile and test its specificity and sensitivity as a biomarker in the diagnosis and prognosis of PaC. METHODS: We obtained serum samples from 197 PaC cases and 158 age- and sex-matched cancer-free controls. We screened the differentially expressed serum miRNAs with Illumina sequencing by synthesis technology using pooled serum samples followed by RT-qPCR validation of a large number of samples arranged in multiple stages. We used risk score analysis to evaluate the diagnostic value of the serum miRNA profiling system. To assess the serum miRNA-based biomarker accuracy in predicting PaC, we performed additional double-blind testing in 77 PaC cases and 52 controls and diagnostic classification in 55 cases with clinically suspected PaC. RESULTS: After the selection and validation process, 7 miRNAs displayed significantly different expression levels in PaC compared with controls. This 7 miRNA-based biomarker had high sensitivity and specificity for distinguishing various stages of PaC from cancer-free controls and also accurately discriminated PaC patients from chronic pancreatitis (CP) patients. Among the 7 miRNAs, miR-21 levels in serum were significantly associated with overall PaC survival. The diagnostic accuracy rate of the 7-miRNA profile was 83.6% in correctly classifying 55 cases with clinically suspected PaC. CONCLUSIONS: These data demonstrate that the 7 miRNA-based biomarker can serve as a novel noninvasive approach for PaC diagnosis and prognosis.

Analysis of clinical characteristics of centrally mediated abdominal pain syndrome, exploration of diagnostic markers and its relationship with the efficacy of duloxetine treatment.

INTRODUCTION: Centrally mediated abdominal pain syndrome (CAPS) is characterized by severe abdominal pain. Diagnosis of CAPS is still an exclusionary diagnosis, there remain no effective diagnostic biomarkers so far. Duloxetine is the major pharmacotherapy of CAPS, while some CAPS patients do not respond to duloxetine treatment. However, there is a lack of molecular markers to predict the efficacy of duloxetine. In our pilot study, we have found differential expression profiles of serum miRNAs between CAPS patients and healthy controls. Our study aims to explore the clinical characteristics, specific miRNAs in serum as diagnostic biomarkers of CAPS and predictive biomarkers of the efficacy of duloxetine. METHODS/DESIGN: In this prospective cohort study, we plan to enroll 430 participants including 215 CAPS patients and 215 healthy controls. The CAPS group takes duloxetine 30 mg per day as an initial dose. Patients will have 24-week medication period and follow up at week 0, 4, 12, 24 and 36. Blood samples will be obtained from patients at every visits and health controls at the initial visit and a series of questionnaires will be completed by the participants. The primary end points are: The differential expression of miRNAs between CAPS groups and healthy control groups at baseline. The changes in abdominal pain scores before and after duloxetine treatment in patients with CAPS and their relationship with the changes in miRNAs. The secondary end point is the changes in scores of depression, anxiety, sleep quality and quality of life before and after duloxetine treatment in patients with CAPS and their relationship with changes in miRNAs. DISCUSSION: Findings of study will provide the reliable basis for diagnosis and the predictor of duloxetine efficacy of CAPS. Importantly, findings grant patients a chance to benefit from treatment.

Pancreatic cancer-derived exosomal microRNA-19a induces ß-cell dysfunction by targeting ADCY1 and EPAC2.

New-onset diabetes mellitus has a rough correlation with pancreatic cancer (PaC), but the underlying mechanism remains unclear. This study aimed to explore the exosomal microRNAs and their potential role in PaC-induced ß-cell dysfunction. The pancreatic ß cells were treated with isolated exosomes from PaC cell lines, SW1990 and BxPC-3, before measuring the glucose-stimulated insulin secretion (GSIS), validating that SW1990 and BxPC-3 might disrupt GSIS of both ß cell line MIN6 and primary mouse pancreatic islets. The difference in expression profiles between exosomes and exosome-free medium of PaC cell lines was further defined, revealing that miR-19a secreted by PaC cells might be an important signaling molecule in this process. Furthermore, adenylyl cyclase 1 (Adcy1) and exchange protein directly activated by cAMP 2 (Epac2) were verified as the direct targets of exogenous miR-19a, which was involved in insulin secretion. These results indicated that exosomes might be an important mediator in the pathogenesis of PaC-DM, and miR-19a might be the effector molecule. The findings shed light on the pathogenesis of PaC-DM.

Upregulation of REG Ialpha accelerates tumor progression in pancreatic cancer with diabetes.

Diabetes is now generally accepted as a crucial event in the process of pancreatic cancer (PaC). However, molecular mechanisms underlying the relationship between diabetes and PaC are not fully understood. Regenerating gene (REG) Ialpha is a growth factor affecting pancreatic islet beta cells, and it has been shown to be involved in the carcinogenesis in gastrointestinal tract. It is rational to speculate that REG Ialpha plays a potential role in the pathogenesis of PaC with diabetes. The aim of this study was to evaluate the REG Ialpha protein expression profile in PaC with and without diabetes, and define the contribution of REG Ialpha on PaC development. We found that REG Ialpha protein preferentially expressed in cancerous tissues of PaC patients with diabetes by Western blot. REG Ialpha positive cancer cells in PaC with diabetes (n = 38) was significantly higher than that in subjects without diabetes (n = 42, p < 0.05) by immunohistochemical analysis. Furthermore, we found that overexpression of REG Ialpha protein in PaC cell lines resulted in accelerated cell proliferation and consequently tumor growth, both in vitro and in vivo. The findings suggest that REG Ialpha may act as one of the tumor promoter and contribute to the aggressive nature of PaC, especially in the subpopulation with diabetes. This study would shed new insights for understanding the molecular mechanisms underlying the link between diabetes and PaC.

XAF1 as a prognostic biomarker and therapeutic target in pancreatic cancer.

XAF1 (X chromosome-linked inhibitor of apoptosis [XIAP]-associated factor 1) is a novel XIAP modulator that negatively regulates the anti-apoptotic effects of XIAP and sensitizes cells to other cell death triggers. It has been reported to be downregulated in a variety of human cancer cell lines. However, the role of XAF1 in pancreatic carcinogenesis remains unclear. In the present study, we investigated the prognostic values of XAF1 expression and its regulation in cancer cell growth and apoptosis both in vitro and in vivo. From the immunohistochemistry staining of tissue microarray, 40 of 89 (44.9%) pancreatic specimens showed low levels of XAF1 expression. Statistical analysis suggested the downregulation of XAF1 was significantly correlated with tumor staging (P = 0.047) and those patients with low XAF1 levels had shorter survival times (P = 0.0162). Multivariate analysis indicated that XAF1 expression was an independent prognostic indicator of the survival of patients with pancreatic cancer (P = 0.007). Furthermore, we found that restoration of XAF1 expression mediated by Ad5/F35 virus suppressed cell proliferation and induced cell cycle arrest and apoptosis, accompanied by the activation of caspases 3, 8, and 9 and poly(ADP-ribose) polymerase as well as increased level of cytochrome c and Bid cleavage. Notably, XAF1 restoration robustly decreased survivin expression rather than XIAP. In addition, in vivo s.c. xenografts from Ad5/F35-XAF1 treatment, which showed less cellular proliferation and enhanced apoptosis, were significantly smaller than those from control groups. Our findings document that XAF1 is a valuable prognostic marker in pancreatic cancer and could be a potential candidate for cancer gene therapy.

Positive expression of L1-CAM is associated with perineural invasion and poor outcome in pancreatic ductal adenocarcinoma.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) frequently invades and migrates along neural tissue, which results in local tumor recurrences, distant metastases, and poor prognosis. We evaluated whether L1 cell adhesion molecule (L1-CAM) and glial cell line-derived neurotrophic factor (GDNF) expression in PDAC correlated with neural invasion and overall survival on a large cohort of previously untreated patients. METHODS: L1-CAM and GDNF were examined by immunohistochemistry in pancreatic cancer tissue samples of 94 cases with PDAC on a tissue microarray. The molecular findings were correlated with pain, clinicopathologic characteristics, and overall survival in these patients. RESULTS: L1-CAM and GDNF were overexpressed in pancreatic cancer tissues compared with the adjacent normal tissues of pancreas. Positive L1-CAM expression was associated with node involvement (P = 0.007), vascular invasion (P = 0.012), perineural invasion (P = 0.001), and higher degree of pain (P = 0.005). In univariate analysis, tissue expression of L1-CAM was associated with poor survival (hazard ratio, 2.508; 95% confidence interval, 1.551-4.053; P < 0.001), and this was also significant in multivariate analysis (hazard ratio, 2.046; 95% confidence interval, 1.200-3.488; P = 0.009). Positive staining of GDNF, neural invasion, and vascular invasion were all statistically significantly related to unfavorable prognosis. CONCLUSIONS: Enhanced expression of L1-CAM may contribute to the pain syndrome and perineural invasion and may correlate with poor overall survival in human pancreatic cancer.

Up-regulation of two actin-associated proteins prompts pulmonary artery smooth muscle cell migration under hypoxia.

Hypoxia stimulates the migration of pulmonary artery smooth muscle cells (PASMCs), which contributes to the pathogenesis of pulmonary vessel structural remodeling in hypoxic pulmonary hypertension (HPH). In the present study, we found, using a proteomics-based method, that gelsolin-like actin-capping protein (CapG) and transgelin were preferentially expressed in human (h)PAMSCs under hypoxia compared with normoxia. These two actin-associated proteins, modulate a variety of physiologic processes, including motility of cells, by interacting differently with the actin cytoskeleton. Our study showed that these two genes were up-regulated at both mRNA and protein levels under hypoxia in hPASMCs. As a key transcriptional regulation factor under hypoxia, hypoxia-inducible factor 1alpha (HIF-1alpha) up-regulated CapG protein expression under normoxia, and knockdown of HIF-1alpha expression in hPASMCs also inhibited hypoxia induced CapG up-regulation. However, HIF-1alpha could not regulate transgelin expression. Reduction of CapG or transgelin expression in hPASMCs by RNA interference was accompanied by significantly impaired migration ability in vitro, especially under hypoxia. Our study demonstrates that CapG and transgelin were preferentially expressed in hPAMSCs under hypoxia compared with normoxia. Hypoxia stimulates expression of these two actin-associated proteins via HIF-1alpha-dependent and -independent pathways, respectively. The up-regulation of these two proteins may contribute to the increased motility of hPASMCs under hypoxia. These findings may contribute to the understanding of the pathogenesis of HPH.