Percutaneous Transhepatic Cholangioscopy and Stone Extraction in a Patient with Recurrent Cholangitis Following Liver Trauma.

Percutaneous transhepatic cholangioscopy (PTCS) is a safe and effective treatment for obstructive biliary stones, when endoscopic retrograde cholangiopancreatography (ERCP) is unsuccessful or unavailable. Once percutaneous access is gained into the biliary tree by an interventional radiologist, the biliary ducts can be directly visualized and any biliary stones can be managed with lithotripsy, mechanical fragmentation, and/or percutaneous extraction. We report a case of a 45-year-old man who sustained a traumatic liver laceration and associated bile duct injury, complicated by bile duct ectasia and intrahepatic biliary stone formation. Despite undergoing a cholecystectomy, multiple ERCPs, and percutaneous transhepatic cholangiogram with drain placement, the underlying problem was not corrected leading to recurrent bouts of gallstone pancreatitis and cholangitis. He was ultimately referred to an interventional radiologist who extracted the impacted intrahepatic biliary stones that were thought to be causing his recurrent infections through cholangioscopy. This is the first case of PTCS with biliary stone extraction in the setting of recurrent biliary obstruction and cholangitis due to traumatic bile duct injury.

First branchial cleft fistula (Work Type 2) with an internal opening to the Eustachian tube: Case report and review of literature.

First branchial cleft anomalies are rare congenital defects of the head and neck. This case report presents a 12-year-old patient with a draining cutaneous pit approximately 1-cm anterior and 5-mm inferior to the right angle of the mandible. Imaging revealed a fistula between the cutaneous pit and Eustachian tube. Further characterization with methylene blue injection into the cutaneous pit resulted in spillage through the right Eustachian tube. Surgical excision of the fistula revealed a cylindrical structure comprised of ectodermal and mesodermal features that most likely represented a Work Type 2 first branchial cleft fistula.

Pervasive introgression facilitated domestication and adaptation in the Bos species complex.

Species of the Bos genus, including taurine cattle, zebu, gayal, gaur, banteng, yak, wisent and bison, have been domesticated at least four times and have been an important source of meat, milk and power for many human cultures. We sequence the genomes of gayal, gaur, banteng, wisent and bison, and provide population genomic sequencing of an additional 98 individuals. We use these data to determine the phylogeny and evolutionary history of these species and show that the threatened gayal is an independent species or subspecies. We show that there has been pronounced introgression among different members of this genus, and that it in many cases has involved genes of considerable adaptive importance. For example, genes under domestication selection in cattle (for example, MITF) were introgressed from domestic cattle to yak. Also, genes in the response-to-hypoxia pathway (for example, EGLN1, EGLN2 and HIF3a) have been introgressed from yak to Tibetan cattle, probably facilitating their adaptation to high altitude. We also validate that there is an association between the introgressed EGLN1 allele and haemoglobin and red blood cell concentration. Our results illustrate the importance of introgression as a source of adaptive variation and during domestication, and suggest that the Bos genus evolves as a complex of genetically interconnected species with shared evolutionary trajectories.

Zinc supplementation for improving glucose handling in pre-diabetes: A double blind randomized placebo controlled pilot study.

AIMS: There are a number of studies showing that zinc supplementation may improve glucose handling in people with established diabetes. We sought to investigate whether this zinc-dependent improvement in glucose handling could potentially be harnessed to prevent the progression of pre-diabetes to diabetes. In this double-blind randomized placebo-controlled trial, we determined participants' fasting blood glucose levels, (FBG) and Homeostasis Model Assessment (HOMA) parameters (beta cell function, insulin sensitivity and insulin resistance) at baseline and after 6 months of zinc supplementation. METHODS: The Bangladesh Institute of Health Sciences Hospital (BIHS) (Mirpur, Dhaka, Bangladesh) database was used to identify 224 patients with prediabetes, of whom 55 met the inclusion criteria and agreed to participate. The participants were randomized either to the intervention or control group using block randomization. The groups received either 30mg zinc sulphate dispersible tablet or placebo, once daily for six months. RESULTS: After six months, the intervention group significantly improved their FBG concentration compared to the placebo group (5.37±0.20mmol/L vs 5.69±0.26, p<0.001) as well as compared to their own baseline (5.37±0.20mmol/L vs 5.8±0.09, p<0.001). Beta cell function, insulin sensitivity and insulin resistance all showed a statistically significant improvement as well. CONCLUSION: To our knowledge this is the first trial to show an improvement in glucose handling using HOMA parameters in participants with prediabetes. Larger randomized controlled trials are warranted to confirm these findings and to explore clinical endpoints.

Identification of Bradyrhizobium elkanii Genes Involved in Incompatibility with Soybean Plants Carrying the Rj4 Allele.

Symbioses between leguminous plants and soil bacteria known as rhizobia are of great importance to agricultural production and nitrogen cycling. While these mutualistic symbioses can involve a wide range of rhizobia, some legumes exhibit incompatibility with specific strains, resulting in ineffective nodulation. The formation of nodules in soybean plants (Glycine max) is controlled by several host genes, which are referred to as Rj genes. The soybean cultivar BARC2 carries the Rj4 gene, which restricts nodulation by specific strains, including Bradyrhizobium elkanii USDA61. Here we employed transposon mutagenesis to identify the genetic locus in USDA61 that determines incompatibility with soybean varieties carrying the Rj4 allele. Introduction of the Tn5 transposon into USDA61 resulted in the formation of nitrogen fixation nodules on the roots of soybean cultivar BARC2 (Rj4 Rj4). Sequencing analysis of the sequence flanking the Tn5 insertion revealed that six genes encoding a putative histidine kinase, transcriptional regulator, DNA-binding transcriptional activator, helix-turn-helix-type transcriptional regulator, phage shock protein, and cysteine protease were disrupted. The cysteine protease mutant had a high degree of similarity with the type 3 effector protein XopD of Xanthomonas campestris. Our findings shed light on the diverse and complicated mechanisms that underlie these highly host-specific interactions and indicate the involvement of a type 3 effector in Rj4 nodulation restriction, suggesting that Rj4 incompatibility is partly mediated by effector-triggered immunity.

SAD-A potentiates glucose-stimulated insulin secretion as a mediator of glucagon-like peptide 1 response in pancreatic ß cells.

Type 2 diabetes is characterized by defective glucose-stimulated insulin secretion (GSIS) from pancreatic ß cells, which can be restored by glucagon-like peptide 1 (GLP-1), an incretin hormone commonly used for the treatment of type 2 diabetes. However, molecular mechanisms by which GLP-1 affects glucose responsiveness in islet ß cells remain poorly understood. Here we investigated a role of SAD-A, an AMP-activated protein kinase (AMPK)-related kinase, in regulating GSIS in mice with conditional SAD-A deletion. We show that selective deletion of SAD-A in pancreas impaired incretin's effect on GSIS, leading to glucose intolerance. Conversely, overexpression of SAD-A significantly enhanced GSIS and further potentiated GLP-1's effect on GSIS from isolated mouse islets. In support of SAD-A as a mediator of incretin response, SAD-A is expressed exclusively in pancreas and brain, the primary targeting tissues of GLP-1 action. Additionally, SAD-A kinase is activated in response to stimulation by GLP-1 through cyclic AMP (cAMP)/Ca(2+)-dependent signaling pathways in islet ß cells. Furthermore, we identified Thr443 as a key autoinhibitory phosphorylation site which mediates SAD-A's effect on incretin response in islet ß cells. Consequently, ablation of Thr443 significantly enhanced GLP-1's effect on GSIS from isolated mouse islets. Together, these findings identified SAD-A kinase as a pancreas-specific mediator of incretin response in islet ß cells.

Synapses of amphids defective (SAD-A) kinase promotes glucose-stimulated insulin secretion through activation of p21-activated kinase (PAK1) in pancreatic ß-Cells.

The p21-activated kinase-1 (PAK1) is implicated in regulation of insulin exocytosis as an effector of Rho GTPases. PAK1 is activated by the onset of glucose-stimulated insulin secretion (GSIS) through phosphorylation of Thr-423, a major activation site by Cdc42 and Rac1. However, the kinase(s) that phosphorylates PAK1 at Thr-423 in islet ß-cells remains elusive. The present studies identified SAD-A (synapses of amphids defective), a member of AMP-activated protein kinase-related kinases exclusively expressed in brain and pancreas, as a key regulator of GSIS through activation of PAK1. We show that SAD-A directly binds to PAK1 through its kinase domain. The interaction is mediated by the p21-binding domain (PBD) of PAK1 and requires both kinases in an active conformation. The binding leads to direct phosphorylation of PAK1 at Thr-423 by SAD-A, triggering the onset of GSIS from islet ß-cells. Consequently, ablation of PAK1 kinase activity or depletion of PAK1 expression completely abolishes the potentiating effect of SAD-A on GSIS. Consistent with its role in regulating GSIS, overexpression of SAD-A in MIN6 islet ß-cells significantly stimulated cytoskeletal remodeling, which is required for insulin exocytosis. Together, the present studies identified a critical role of SAD-A in the activation of PAK1 during the onset of insulin exocytosis.

The burden of multidrug-resistant organisms on tertiary hospitals posed by patients with recent stays in long-term acute care facilities.

BACKGROUND: Long-term acute care (LTAC) facilities admit patients with complex, advanced disease states. Study aims were to determine the burden posed on hospitals associated with LTAC exposure and analyze the differences between "present on admission" (POA) multidrug-resistant (MDR), gram-negative organisms (GNO) and POA MDR gram-positive organisms (GPO). METHODS: A multicenter retrospective study was conducted in 13 hospitals from southeast Michigan, from September 1, 2008, to August 31, 2009. Cultures obtained in the first 72 hours of hospitalization (ie, POA) of MDR-GPO and MDR-GNO were reviewed. LTAC exposures in the previous 6 months and direct admission from a LTAC were recorded. RESULTS: Overall, 5,297 patients with 7,147 MDR POA cultures were analyzed: 2,619 (36.6%) were MDR-GNO, and 4,528 (63.4%) were MDR-GPO. LTAC exposure in the past 6 months was present in 251 (5.2%) infectious episodes and was significantly more common among POA MDR-GNO than MDR-GPO (158 [8.6%] and 94 [3.1%], respectively, odds ratio, 2.87; P < .001). Recent LTAC exposure was strongly associated with both carbapenem-resistant Enterobacteriaceae (CRE) (31.6% of all CRE cases, P < .001) and Acinetobacter baumannii (14.9% of all A baumannii cases, P < .001). CONCLUSION: Nearly 10% of MDR-GNO POA had recent LTAC exposure. Hospital efforts to control the spread of MDR-GNO should focus on collaborations and communications with referring LTACs and interventions targeted towards patients with recent LTAC exposure.

Cell-permeable peptide-based disruption of endogenous PKA-AKAP complexes: a tool for studying the molecular roles of AKAP-mediated PKA subcellular anchoring.

Stimulation of numerous G protein-coupled receptors leads to the elevation of intracellular concentrations of cAMP, which subsequently activates the PKA pathway. Specificity of the PKA signaling module is determined by a sophisticated subcellular targeting network that directs the spatiotemporal activation of the kinase. This specific compartmentalization mechanism occurs through high-affinity interactions of PKA with A-kinase anchoring proteins (AKAPs), the role of which is to target the kinase to discrete subcellular microdomains. Recently, a peptide designated "AKAPis" has been proposed to competitively inhibit PKA-AKAP interactions in vitro. We therefore sought to characterize a cell-permeable construct of the AKAPis inhibitor and use it as a tool to characterize the impact of PKA compartmentalization by AKAPs. Using insulin-secreting pancreatic beta-cells (INS-1 cells), we showed that TAT-AKAPis (at a micromolar range) dose dependently disrupted a significant fraction of endogenous PKA-AKAP interactions. Immunoflurescent analysis also indicated that TAT-AKAPis significantly affected PKA subcellular localization. Furthermore, TAT-AKAPis markedly attenuated glucagon-induced phosphorylations of p44/p42 MAPKs and cAMP response element binding protein, which are downstream effectors of PKA. In parallel, TAT-AKAPis dose dependently inhibited the glucagon-induced potentiation of insulin release. Therefore, AKAP-mediated subcellular compartmentalization of PKA represents a key mechanism for PKA-dependent phosphorylation events and potentiation of insulin secretion in intact pancreatic beta-cells. More interestingly, our data highlight the effectiveness of the cell-permeable peptide-mediated approach to monitoring in cellulo PKA-AKAP interactions and delineating PKA-dependent phosphorylation events underlying specific cellular responses.

Glucagon promotes cAMP-response element-binding protein phosphorylation via activation of ERK1/2 in MIN6 cell line and isolated islets of Langerhans.

By using the MIN6 cell line and pancreatic islets, we show that in the presence of a low glucose concentration, corresponding to physiological glucagon release from alpha cells, glucagon treatment of the beta cell caused a rapid, time-dependent phosphorylation and activation of p44/p42 mitogen-activated protein kinase (ERK1/2) independently from extracellular calcium influx. Inhibition of either cAMP-dependent protein kinase (PKA) or MEK completely blocked ERK1/2 activation by glucagon. However, no significant activation of several upstream activators of MEK, including Shc-p21(Ras) and phosphatidylinositol 3-kinase, was observed in response to glucagon treatment. Chelation of intracellular calcium (intracellular [Ca(2+)]) reduced glucagon-mediated ERK1/2 activation. In addition, internalization of glucagon receptors through clathrin-coated pits formation is required for ERK1/2 activation. Remarkably, glucagon promotes the nuclear translocation of ERK1/2 and induces the phosphorylation of cAMP-response element-binding protein (CREB). Miniglucagon, produced from glucagon and released together with the mother hormone from the alpha cells in low glucose situations, blocks the insulinotropic effect of glucagon, whereas it does not inhibit the glucagon-induced PKA/ERK1/2/CREB pathway. We conclude that glucagon-induced ERK1/2 activation is mediated by PKA and that an increase in [Ca(2+)](i) is required for maximal ERK activation. Our results uncover a novel mechanism by which the PKA/ERK1/2 signaling network engaged by glucagon, in situation of low glucose concentration, regulates phosphorylation of CREB, a transcription factor crucial for normal beta cell function and survival.

Cooperative effects between protein kinase A and p44/p42 mitogen-activated protein kinase to promote cAMP-responsive element binding protein activation after beta cell stimulation by glucose and its alteration due to glucotoxicity.

Long-term hyperglycemia, a major characteristic of the diabetic state, contributes to the deterioration of the beta cell function, a concept known as beta cell glucotoxicity. We used the MIN6 beta cell line and isolated rat islets to clarify the signaling mechanism(s) used by glucose to activate cAMP-responsive element binding protein (CREB), a transcription factor crucial for beta cell biology, and to evaluate the possible downregulation of this mechanism mediated by long-term hyperglycemia. We report that glucose (10 mM) induces an increase in cytosolic calcium concentration that leads to cAMP-induced protein kinase A (PKA) activation, promoting nuclear translocation of activated ERK1/2. The observation that glucose-induced CREB phosphorylation was totally inhibited by the PKA inhibitor H89 (2 microM) and reduced by 50% with the ERK1/2 inhibitor PD98059 (20 microM) indicates that ERK1/2, located downstream of PKA, cooperates with PKA and is responsible for half of the PKA-mediated CREB phosphorylation elicited by glucose in MIN6 beta cells. We also found that exposure of mu cells for 24 h to high glucose (25 mM) induced a 70% decrease in cellular ERK1/2 and a 50% decrease in CREB content. In high-glucose-treated, ERK1/2- and CREB-downregulated beta cells, there was a loss of glucose (10 mM, 5 min)-stimulated ERK1/2 and CREB phosphorylation that was associated with nuclear apoptotic characteristics. Since we have shown that activation of ERK1/2 is crucial for CREB phosphorylation, loss of the ERK1/2-CREB signaling pathway in beta cells due to long-term hyperglycemia is likely to exacerbate beta cell failure in diabetic states by affecting physiologically relevant gene expression and by inducing apoptosis.