Knockout of the Tachykinin Receptor 1 in the Mdr2-/- (Abcb4-/-) Mouse Model of Primary Sclerosing Cholangitis Reduces Biliary Damage and Liver Fibrosis.

Activation of the substance P (SP)/neurokinin 1 receptor (NK1R) axis triggers biliary damage/senescence and liver fibrosis in bile duct ligated and Mdr2-/- (alias Abcb4-/-) mice through enhanced transforming growth factor-ß1 (TGF-ß1) biliary secretion. Recent evidence indicates a role for miR-31 (MIR31) in TGF-ß1-induced liver fibrosis. We aimed to define the role of the SP/NK1R/TGF-ß1/miR-31 axis in regulating biliary proliferation and liver fibrosis during cholestasis. Thus, we generated a novel model with double knockout of Mdr2-/- and NK1R-/ (alias Tacr1-/-) to further address the role of the SP/NK1R axis during chronic cholestasis. In vivo studies were performed in the following 12-week-old male mice: (i) NK1R-/-; (ii) Mdr2-/-; and (iii) NK1R-/-/Mdr2-/- (Tacr1-/-/Abcb4-/-) and their corresponding wild-type controls. Liver tissues and cholangiocytes were collected, and liver damage, changes in biliary mass/senescence, and inflammation as well as liver fibrosis were evaluated by both immunohistochemistry in liver sections and real-time PCR. miR-31 expression was measured by real-time PCR in isolated cholangiocytes. Decreased ductular reaction, liver fibrosis, biliary senescence, and biliary inflammation were observed in NK1R-/-/Mdr2-/- mice compared with Mdr2-/- mice. Elevated expression of miR-31 was observed in Mdr2-/- mice, which was reduced in NK1R-/-/Mdr2-/- mice. Targeting the SP/NK1R and/or miR-31 may be a potential approach in treating human cholangiopathies, including primary sclerosing cholangitis.

The Secretin/Secretin Receptor Axis Modulates Ductular Reaction and Liver Fibrosis through Changes in Transforming Growth Factor-ß1-Mediated Biliary Senescence.

Activation of the secretin (Sct)/secretin receptor (SR) axis stimulates ductular reaction and liver fibrosis, which are hallmarks of cholangiopathies. Our aim was to define the role of Sct-regulated cellular senescence, and we demonstrated that both ductular reaction and liver fibrosis are significantly reduced in Sct-/-, SR-/-, and Sct-/-/SR-/- bile duct ligated (BDL) mice compared with BDL wild-type mice. The reduction in hepatic fibrosis in Sct-/-, SR-/-, and Sct-/-/SR-/- BDL mice was accompanied by reduced transforming growth factor-ß1 levels in serum and cholangiocyte supernatant, as well as decreased expression of markers of cellular senescence in cholangiocytes in contrast to enhanced cellular senescence in hepatic stellate cells compared with BDL wild-type mice. Secretin directly stimulated the senescence of cholangiocytes and regulated, by a paracrine mechanism, the senescence of hepatic stellate cells and liver fibrosis via modulation of transforming growth factor-ß1 biliary secretion. Targeting senescent cholangiocytes may represent a novel therapeutic approach for ameliorating hepatic fibrosis during cholestatic liver injury.

Knockout of secretin receptor reduces biliary damage and liver fibrosis in Mdr2-/- mice by diminishing senescence of cholangiocytes.

Secretin receptor (SR), only expressed by cholangiocytes, plays a key role in the regulation of biliary damage and liver fibrosis. The aim of this study was to determine the effects of genetic depletion of SR in Mdr2-/- mice on intrahepatic biliary mass, liver fibrosis, senescence, and angiogenesis. 12 wk SR-/-, Mdr2-/-, and SR-/-/Mdr2-/- mice with corresponding wild-type mice were used for the in vivo studies. Immunohistochemistry or immunofluorescence was performed in liver sections for (i) biliary expression of SR; (ii) hematoxylin and eosin; (iii) intrahepatic biliary mass by CK-19; (iv) fibrosis by Col1a1 and α-SMA; (v) senescence by SA-ß-gal and p16; and (vi) angiogenesis by VEGF-A and CD31. Secretin (Sct) and TGF-ß1 levels were measured in serum and cholangiocyte supernatant by ELISA. In total liver, isolated cholangiocytes or HSCs, we evaluated the expression of fibrosis markers (FN-1 and Col1a1); senescence markers (p16 and CCL2); microRNA 125b and angiogenesis markers (VEGF-A, VEGFR-2, CD31, and vWF) by immunoblots and/or qPCR. In vitro, we measured the paracrine effect of cholangiocyte supernatant on the expression of senescent and fibrosis markers in human hepatic stellate cells (HHSteCs). The increased level of ductular reaction, fibrosis, and angiogenesis in Mdr2-/- mice was reduced in SR-/-/Mdr2-/- mice. Enhanced senescence levels in cholangiocytes from Mdr2-/- mice were reversed to normal in SR-/-/Mdr2-/- mice. However, senescence was decreased in HSCs from Mdr2-/- mice but returned to normal values in SR-/-/Mdr2-/- mice. In vitro treatment of HHSteCs with supernatant from cholangiocyte lacking SR (containing lower biliary levels of Sct-dependent TGF-ß1) have decreased fibrotic reaction and increased cellular senescence. Sct-induced TGF-ß1 secretion was mediated by microRNA 125b. Our data suggest that differential modulation of angiogenesis-dependent senescence of cholangiocytes and HSCs may be important for the treatment of liver fibrosis in cholangiopathies.

Mechanisms of cholangiocyte responses to injury.

Cholangiocytes, epithelial cells that line the biliary epithelium, are the primary target cells for cholangiopathies including primary sclerosing cholangitis and primary biliary cholangitis. Quiescent cholangiocytes respond to biliary damage and acquire an activated neuroendocrine phenotype to maintain the homeostasis of the liver. The typical response of cholangiocytes is proliferation leading to bile duct hyperplasia, which is a characteristic of cholestatic liver diseases. Current studies have identified various signaling pathways that are associated with cholangiocyte proliferation/loss and liver fibrosis in cholangiopathies using human samples and rodent models. Although recent studies have demonstrated that extracellular vesicles and microRNAs could be mediators that regulate these messenger/receptor axes, further studies are required to confirm their roles. This review summarizes current studies of biliary response and cholangiocyte proliferation during cholestatic liver injury with particular emphasis on the secretin/secretin receptor axis. This article is part of a Special Issue entitled: Cholangiocytes in Health and Diseaseedited by Jesus Banales, Marco Marzioni, Nicholas LaRusso and Peter Jansen.

Melatonin inhibits hypothalamic gonadotropin-releasing hormone release and reduces biliary hyperplasia and fibrosis in cholestatic rats.

Melatonin is a hormone produced by the pineal gland with increased circulating levels shown to inhibit biliary hyperplasia and fibrosis during cholestatic liver injury. Melatonin also has the capability to suppress the release of hypothalamic gonadotropin-releasing hormone (GnRH), a hormone that promotes cholangiocyte proliferation when serum levels are elevated. However, the interplay and contribution of neural melatonin and GnRH to cholangiocyte proliferation and fibrosis in bile duct-ligated (BDL) rats have not been investigated. To test this, cranial levels of melatonin were increased by implanting osmotic minipumps that performed an intracerebroventricular (ICV) infusion of melatonin or saline for 7 days starting at the time of BDL. Hypothalamic GnRH mRNA and cholangiocyte secretion of GnRH and melatonin were assessed. Cholangiocyte proliferation and fibrosis were measured. Primary human hepatic stellate cells (HSCs) were treated with cholangiocyte supernatants, GnRH, or the GnRH receptor antagonist cetrorelix acetate, and cell proliferation and fibrosis gene expression were assessed. Melatonin infusion reduced hypothalamic GnRH mRNA expression and led to decreased GnRH and increased melatonin secretion from cholangiocytes. Infusion of melatonin was found to reduce hepatic injury, cholangiocyte proliferation, and fibrosis during BDL-induced liver injury. HSCs supplemented with BDL cholangiocyte supernatant had increased proliferation, and this increase was reversed when HSCs were supplemented with supernatants from melatonin-infused rats. GnRH stimulated fibrosis gene expression in HSCs, and this was reversed by cetrorelix acetate cotreatment. Increasing bioavailability of melatonin in the brain may improve outcomes during cholestatic liver disease.NEW & NOTEWORTHY We have previously demonstrated that GnRH is expressed in cholangiocytes and promotes their proliferation during cholestasis. In addition, dark therapy, which increases melatonin, reduced cholangiocyte proliferation and fibrosis during cholestasis. This study expands these findings by investigating neural GnRH regulation by melatonin during BDL-induced cholestasis by infusing melatonin into the brain. Melatonin infusion reduced cholangiocyte proliferation and fibrosis, and these effects are due to GNRH receptor 1-dependent paracrine signaling between cholangiocytes and hepatic stellate cells.

Idioventricular Rhythm: A Rare Presentation of Left Ventricular Pseudoaneurysm Following Radiofrequency Ablation.

Left ventricular pseudoaneurysm (PsA) is a rare complication of radiofrequency ablation (RFA) of cardiac arrhythmias. Presentation can vary widely in terms of timeline, signs, and symptoms. Idioventricular rhythm is a rare presentation of PsA post-ablation. No cases of post-ablation PsA presenting with idioventricular rhythm have been reported in the literature to date. A 72-year-old male presented with symptomatic idioventricular rhythm 34 days post RFA for premature ventricular complexes (PVCs). A PsA involving the distal anterolateral of his left ventricle wall was identified on transthoracic echo and computed tomography (CT). This patient underwent surgical patch repair which resolved his ventricular arrhythmia.

Primary Peritoneal Mesothelioma Affecting the Greater Omentum That Mimicked an Omental Infarction: A Case Report.

Introduction: Malignant peritoneal mesothelioma (MPM) is a rare cancer that is associated with asbestos exposure. The diagnosis can be difficult given the nonspecific nature of presenting symptoms and the presence of concomitant confounding findings. Case Presentation: We report a 71-year-old male who presented with right lower quadrant pain and new-onset ascites. CT imaging of the abdomen/pelvis demonstrated omental stranding concerning for a possible omental infarction. Subsequent imaging showed persistent omental edema but no identifiable soft tissue mass. A biopsy of the omentum showed atypical mesothelial proliferation, but pathology was unable to determine if proliferation was a neoplastic versus reactive process. Surgical oncology performed a diagnostic laparoscopy that showed peritoneal studding of the omentum. Subsequent immunohistochemical staining of the omentum demonstrated preservation of BAP1 expression and loss of MTAP expression, consistent with peritoneal mesothelioma. Conclusion: MPM is a rare and aggressive cancer with an overall poor prognosis. The diagnosis of MPM can be difficult based on the nonspecific clinical presentation, insufficient imaging and laboratory testing, and the presence of concomitant confounding findings, such as with this patient and his admitting diagnosis of omental infarction. This case demonstrates the importance of developing a broad differential while maintaining an awareness of heuristics that can influence clinical decision-making.

Associations of the microbiome and esophageal disease.

The incidence of esophageal diseases such as esophageal adenocarcinoma (EAC) and gastroesophageal reflux disease (GERD) have been increasing over the last 40 years. The esophageal microbiome appears to have a role in the development of some disease processes, and could also serve as markers of early diseases of the esophagus. A literature review was performed examining the role of the microbiome in the development of esophageal disease. In addition, the results of several studies and experiments were included in the review. Both EAC and GERD have increased in incidence over the last 40 years. Barrett's esophagus (BE) is a risk factor for EAC. Patients with BE appear to have a microbiome expression pattern distinct from patients without BE. The distinct pattern may be related to factors within the distal esophagus such as a more acidic environment, intraluminal stasis and other elements. It remains unclear whether the change in microflora leads to esophageal disease, or whether the disease process within the esophagus allows these particular organisms to experience overgrowth compared to other microflora. Patient factors such as body mass index (BMI), diet and geographic location also appear to affect the esophageal microbiome. There is an association with the esophageal microbiome and several esophageal diseases. Future studies should examine these correlations more closely. The distinct patterns may be able to serve as a marker of early disease, and possibly lead to a mechanism for the development of esophageal disease.

Author Correction: The role of the secretin/secretin receptor axis in inflammatory cholangiocyte communication via extracellular vesicles.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

The role of the secretin/secretin receptor axis in inflammatory cholangiocyte communication via extracellular vesicles.

Small and large intrahepatic bile ducts consist of small and large cholangiocytes, respectively, and these cholangiocytes have different morphology and functions. The gastrointestinal peptide hormone, secretin (SCT) that binds to secretin receptor (SR), is a key mediator in cholangiocyte pathophysiology. Extracellular vesicles (EVs) are membrane-bound vesicles and cell-cell EV communication is recognized as an important factor in liver pathology, although EV communication between cholangiocytes is not identified to date. Cholangiocytes secrete proinflammatory cytokines during bacterial infection leading to biliary inflammation and hyperplasia. We demonstrate that cholangiocytes stimulated with lipopolysaccharide (LPS), which is a membrane component of gram-negative bacteria, secrete more EVs than cholangiocytes incubated with vehicle. These LPS-derived EVs induce inflammatory responses in other cholangiocytes including elevated cytokine production and cell proliferation. Large but not small cholangiocytes show inflammatory responses against large but not small cholangiocyte-derived EVs. Large cholangiocytes with knocked down either SCT or SR by short hairpin RNAs show reduced EV secretion during LPS stimulation, and EVs isolated from SCT or SR knocked down cholangiocytes fail to induce inflammatory reactions in control large cholangiocytes. This study identifies cholangiocyte EV communication during LPS stimulation, and demonstrates that the SCT/SR axis may be important for this event.

Gender difference in academic planning activity among medical students.

BACKGROUND: In Vietnam, as doctor of medicine is socially considered a special career, both men and women who are enrolled in medical universities often study topics of medicine seriously. However, as culturally expected, women often perform better than men. Because of this, teaching leadership and management skill (LMS) to develop academic planning activity (APA) for female medical students would also be expected to be more effective than male counterparts. This research aimed to compare by gender the effect of teaching LMS on increasing APA, using propensity score matching (PSM). METHODS: In a cross-sectional survey utilizing a self-reported structured questionnaire on a systematic random sample of 421 male and female medical students in Hanoi Medical University, this study adopted first regression techniques to construct a fit model, then PSM to create a matched control group in order to allow for evaluating the effect of LMS education. RESULTS: There were several interesting gender differences. First, while for females LMS education had both direct and indirect effects on APA, it had only direct effect on males' APA. Second, after PSM to adjust for the possible confounders to balance statistically two groups - with and without LMS education, there is statistically a significant difference in APA between male and female students, making a net difference of 11% (p<.01), equivalent to 173 students. The difference in APA between exposed and matched control group in males and females was 9% and 20%, respectively. These estimates of 9.0 and 20.0 percentage point increase can be translated into the practice of APA by 142 males and 315 females, respectively, in the population. These numbers of APA among male and female students can be explained by LMS education. CONCLUSIONS: Gender appears to be a factor explaining in part academic planning activity.