Structure of lipoprotein lipase in complex with GPIHBP1.

Lipoprotein lipase (LPL) plays a central role in triglyceride (TG) metabolism. By catalyzing the hydrolysis of TGs present in TG-rich lipoproteins (TRLs), LPL facilitates TG utilization and regulates circulating TG and TRL concentrations. Until very recently, structural information for LPL was limited to homology models, presumably due to the propensity of LPL to unfold and aggregate. By coexpressing LPL with a soluble variant of its accessory protein glycosylphosphatidylinositol-anchored high-density lipoprotein binding protein 1 (GPIHBP1) and with its chaperone protein lipase maturation factor 1 (LMF1), we obtained a stable and homogenous LPL/GPIHBP1 complex that was suitable for structure determination. We report here X-ray crystal structures of human LPL in complex with human GPIHBP1 at 2.5-3.0 Å resolution, including a structure with a novel inhibitor bound to LPL. Binding of the inhibitor resulted in ordering of the LPL lid and lipid-binding regions and thus enabled determination of the first crystal structure of LPL that includes these important regions of the protein. It was assumed for many years that LPL was only active as a homodimer. The structures and additional biochemical data reported here are consistent with a new report that LPL, in complex with GPIHBP1, can be active as a monomeric 1:1 complex. The crystal structures illuminate the structural basis for LPL-mediated TRL lipolysis as well as LPL stabilization and transport by GPIHBP1.

A Clinical Report Utilizing the VITOM 3D® Microvideoscope for Cleft Palate Repair.

Cleft palate surgery has traditionally presented numerous problems for cleft surgeons including ergonomics, limited visual fields restricting the opportunity for demonstration and teaching. Additionally, the move toward online teaching means the ability to record or livestream video is paramount. The following report of eight cleft palate repairs highlights the novel use of the Vitom 3D® microvideoscope as an innovative technique for cleft palate repair with our early experience demonstrating significant ergonomic and teaching benefits.

Performance measures for ERCP and endoscopic ultrasound: a European Society of Gastrointestinal Endoscopy (ESGE) Quality Improvement Initiative.

The European Society of Gastrointestinal Endoscopy and United European Gastroenterology present a short list of key performance measures for endoscopic ultrasound (EUS) and endoscopic retrograde cholangiopancreatography (ERCP). We recommend that endoscopy services across Europe adopt the following seven key and one minor performance measures for EUS and ERCP, for measurement and evaluation in daily practice at center and endoscopist level: 1: Adequate antibiotic prophylaxis before ERCP (key performance measure, at least 90 %); 2: Antibiotic prophylaxis before EUS-guided puncture of cystic lesions (key performance measure, at least 95 %); 3: Bile duct cannulation rate (key performance measure, at least 90 %); 4: Tissue sampling during EUS (key performance measure, at least 85 %); 5: Appropriate stent placement in patients with biliary obstruction below the hilum (key performance measure, at least 95 %); 6: Bile duct stone extraction (key performance measure, at least 90 %); 7: Post-ERCP pancreatitis (key performance measure, less than 10 %). 8: Adequate documentation of EUS landmarks (minor performance measure, at least 90 %).This present list of quality performance measures for ERCP and EUS recommended by ESGE should not be considered to be exhaustive: it might be extended in future to address further clinical and scientific issues.

The Effects of Twelve Weeks of Tai Chi Practice on Anxiety in Stressed But Healthy People Compared to Exercise and Wait-List Groups-A Randomized Controlled Trial.

OBJECTIVE: This randomized controlled trial was undertaken to determine whether 12 weeks of Tai Chi (TC) practice can reduce anxiety in healthy but stressed people. METHOD: Fifty participants were randomized into TC (n=17), exercise (n=17), and wait-list (WL) groups (n=16). Outcome measures used were State Trait Anxiety Inventory, Perceived Stress Scale 14 (PSS14), blood pressure and heart rate variability, visual analogue scale (VAS), and Short Form 36. RESULTS: Significant improvements were observed from baseline for both TC and exercise groups for both state (p <0.01) and trait (p <0.01) anxiety, PSS14 (p <0.01), VAS (p <0.01), mental health domain (p <0.01), and vitality domain (p <0.01). Superior outcomes were also observed for TC when compared with WL for state and trait anxiety (p <0.01) and mental health domain (p <0.05). CONCLUSION: TC reduces stress levels in healthy individuals and provides a safer, cost effective, and less physically vigorous alternative to exercise.

Update in Natural Orifice Translumenal Endoscopic Surgery (NOTES).

PURPOSE OF REVIEW: The recent developments and clinical applications of natural orifice translumenal endoscopic surgery (NOTES)-procedures and technologies are going to be presented. RECENT FINDINGS: In experimental as well as clinical settings, NOTES-procedures are predominantly performed in hybrid technique. Current experimental studies focus on the implementation of new surgical approaches as well as on the training of procedures. One emphasis in the clinical application is transrectal and transanal interventions. Transanal total mesorectal excision is equivalent to laparoscopic procedures but with the benefit of an even less invasive access. Transvaginal cholecystectomy can achieve results that are comparable to surgeries that are performed with laparoscopic techniques alone. An analysis of the German NOTES-Register concerning appendectomies as well as the national performance of NOTES-interventions in Switzerland is presented. Apart from intraabdominal approaches, several centers proclaim transoral thyroidectomies and transoral mediastinoscopies. SUMMARY: NOTES-procedures are performed in animal experiments as well as in clinical setting although with less frequency. At this time, hybrid techniques using rigid instruments are mainly applied.

[Update in pediatric endoscopy]. / Pädiatrische Endoskopie ­ ein Update.

Pediatric endoscopy is a good example of a multidiscipliniary approach. In many, but not all hospitals, endoscopy in pediatric patients is performed by conventional gastroenterologists and only in some centers by pediatric gastroenterologists. This is due to the fact that not as many pediatric gastroenterologists are available. Some of the centers are very experienced. There is only limited literature on pediatric endoscopy available. Therefore, an update may be relevant for both the adult and pediatric gastroenterologists. Here we describe current knowledge on endoscopic procedures in pediatric patients.

Differential cellular expression of organic anion transporting peptides OATP1A2 and OATP2B1 in the human retina and brain: implications for carrier-mediated transport of neuropeptides and neurosteriods in the CNS.

Organic anion transporting polypeptides (OATPs) are polyspecific organic anion transporters, which are expressed in the blood-brain barrier, the choroid plexus, and other organs. The physiologic function of OATPs in extrahepatic tissues remains ambiguous. In rat retina, members of the OATP family are expressed. We therefore investigated the human retina for the expression of OATP1A2 and OATP2B1 and extended the study to human brain. Furthermore, we searched for peptide neurotransmitters as novel OATP substrates. OATP1A2 displayed a broad expression pattern in human retina as assessed by immunofluorescence localization. It is expressed in photoreceptor bodies and somas of amacrine cells. OATP1B2 expression is restricted to the inner nuclear layer and to the inner plexiform layer. Using paraffin sections from human cortex, cerebellum, and hippocampus, OATP1A2 was localized to neurons and neuronal processes, while OATP2B1 is expressed in endothelial cells of brain capillaries. Substance P and vasoactive intestinal peptide were identified as substrates for OATP1A2 and OATP2B1. Double-labeling immunofluorescence of human retina demonstrated the presence of substance P and of vasoactive intestinal peptides in neurons expressing OATP1A2 and OATP2B1, respectively. The expression of OATP1A2 and OATP2B1 in retinal neurons implies a role of these transporters in the reuptake of peptide neurotransmitters released from retinal neurons. The abundant expression of OATP1A2 in brain neurons points to the possibility that OATP1A2 could be involved in the homeostasis of neurosteroids. The high expression of OATP2B1 in brain capillaries supports an important function of OATPs in substance penetration across the blood-brain barrier.

Ligandability Assessment of IL-1ß by Integrated Hit Identification Approaches.

Human interleukin-1ß (IL-1ß) is a pro-inflammatory cytokine that plays a critical role in the regulation of the immune response and the development of various inflammatory diseases. In this publication, we disclose our efforts toward the discovery of IL-1ß binders that interfere with IL-1ß signaling. To this end, several technologies were used in parallel, including fragment-based screening (FBS), DNA-encoded library (DEL) technology, peptide discovery platform (PDP), and virtual screening. The utilization of distinct technologies resulted in the identification of new chemical entities exploiting three different sites on IL-1ß, all of them also inhibiting the interaction with the IL-1R1 receptor. Moreover, we identified lysine 103 of IL-1ß as a target residue suitable for the development of covalent, low-molecular-weight IL-1ß antagonists.

Detecting S-adenosyl-L-methionine-induced conformational change of a histone methyltransferase using a homogeneous time-resolved fluorescence-based binding assay.

A homogeneous time-resolved fluorescence (HTRF)-based binding assay has been established to measure the binding of the histone methyltransferase (HMT) G9a to its inhibitor CJP702 (a biotin analog of the known peptide-pocket inhibitor, BIX-01294). This assay was used to characterize G9a inhibitors. As expected, the peptide-pocket inhibitors decreased the G9a-CJP702 binding signal in a concentration-dependent manner. In contrast, the S-adenosyl-L-methionine (SAM)-pocket compounds, SAM and sinefungin, significantly increased the G9a-CJP702 binding signal, whereas S-adenosyl-L-homocysteine (SAH) showed minimal effect. Enzyme kinetic studies showed that CJP702 is an uncompetitive inhibitor (vs. SAM) that has a strong preference for the E:SAM form of the enzyme. Other data presented suggest that the SAM/sinefungin-induced increase in the HTRF signal is secondary to an increased E:SAM or E:sinefungin concentration. Thus, the G9a-CJP702 binding assay not only can be used to characterize the peptide-pocket inhibitors but also can detect the subtle conformational differences induced by the binding of different SAM-pocket compounds. To our knowledge, this is the first demonstration of using an uncompetitive inhibitor as a probe to monitor the conformational change induced by compound binding with an HTRF assay.

Probing the bioactivity-relevant chemical space of robust reactions and common molecular building blocks.

In the search for new bioactive compounds, there is a trend toward increasingly complex compound libraries aiming to target the demanding targets of the future. In contrast, medicinal chemistry and traditional library design rely mainly on a small set of highly established and robust reactions. Here, we probe a set of 58 such reactions for their ability to sample the chemical space of known bioactive molecules, and the potential to create new scaffolds. Combined with ~26,000 common available building blocks, the reactions retrieve around 9% of a scaffold-diverse set of compounds active on human target proteins covering all major pharmaceutical target classes. Almost 80% of generated scaffolds from virtual one-step synthesis products are not present in a large set of known bioactive molecules for human targets, indicating potential for new discoveries. The results suggest that established synthesis resources are well suited to cover the known bioactivity-relevant chemical space and that there are plenty of unexplored regions accessible by these reactions, possibly providing valuable "low-hanging fruit" for hit discovery.

A collection of robust organic synthesis reactions for in silico molecule design.

A focused collection of organic synthesis reactions for computer-based molecule construction is presented. It is inspired by real-world chemistry and has been compiled in close collaboration with medicinal chemists to achieve high practical relevance. Virtual molecules assembled from existing starting material connected by these reactions are supposed to have an enhanced chance to be amenable to real chemical synthesis. About 50% of the reactions in the dataset are ring-forming reactions, which fosters the assembly of novel ring systems and innovative chemotypes. A comparison with a recent survey of the reactions used in early drug discovery revealed considerable overlaps with the collection presented here. The dataset is available encoded as computer-readable Reaction SMARTS expressions from the Supporting Information presented for this paper.

Genetic determinants of drug-induced cholestasis and intrahepatic cholestasis of pregnancy.

Intrahepatic cholestasis of pregnancy and drug-induced cholestasis are two clinically important forms of acquired cholestatic liver disease. The understanding of the underlying mechanisms of acquired cholestasis has recently made considerable progress by the identification of canalicular ATP-binding cassette (ABC) transporters as likely targets for these forms of cholestasis. Cholestasis of pregnancy is linked to estrogen and progesterone metabolites. These metabolites have been shown to impair the bile salt export pump (BSEP) function by an indirect mechanism. In addition, genetic variants (as well as mutants) of the genes coding for the phosphatidylcholine translocator MDR3 and BSEP and for the farnesoid X receptor, which is critical in the transcriptional activation of MDR3 ( ABCB4) and BSEP ( ABCB11) have been associated with intrahepatic cholestasis of pregnancy. The pathogenesis of drug-induced liver injury encompasses a wide spectrum of mechanisms, some of which are still poorly understood. BSEP is now known to be subject to drug inhibition in susceptible patients. Information on genetic factors rendering individuals susceptible to inhibition of BSEP by drugs or their metabolites is still scarce. Besides rare mutations that have been linked to drug-induced cholestasis, the common p.V444A polymorphism of BSEP has been identified as a potential risk factor. In this review, the authors summarize key concepts of physiology of bile formation, diagnostic principles to indentify these forms of acquired cholestasis, as well as pathogenetic mechanisms leading to intrahepatic cholestasis of pregnancy or drug-induced cholestasis. In addition, they review the current knowledge on genetic susceptibility factors for these two forms of cholestasis.

ABC-transporters are localized in caveolin-1-positive and reggie-1-negative and reggie-2-negative microdomains of the canalicular membrane in rat hepatocytes.

UNLABELLED: The canalicular plasma membrane is constantly exposed to bile acids acting as detergents. Bile acids are essential to mediate release of biliary lipids from the canalicular membrane. Membrane microdomains (previously called lipid rafts) are biochemically defined by their resistance to detergent solubilization at cold temperature. We aimed to investigate the canalicular plasma membrane for the presence of microdomains, which could protect this membrane against the detergent action of bile acids. Highly purified rat liver canalicular plasma membrane vesicles were extracted with 1% Triton X-100 or 1% Lubrol WX at 4 degrees C and subjected to flotation through sucrose step gradients. Both detergents yielded detergent-resistant membranes containing the microdomain markers alkaline phosphatase and sphingomyelin. However, cholesterol was resistant to Lubrol WX solubilization, whereas it was only marginally resistant to solubilization by Triton X-100. The microdomain marker caveolin-1 was localized to the canalicular plasma membrane domain and was resistant to Lubrol WX, but to a large extent solubilized by Triton X-100. The two additional microdomain markers, reggie-1 and reggie-2, were localized to the basolateral and canalicular plasma membrane and were partially resistant to Lubrol WX but resistant to Triton X-100. The canalicular transporters bile salt export pump, multidrug resistance protein 2, multidrug resistance-associated protein 2, and Abcg5 were largely resistant to Lubrol WX but were solubilized by Triton X-100. CONCLUSION: These results indicate the presence of two different types of microdomains in the canalicular plasma membrane: "Lubrol-microdomains" and "Triton-microdomains". "Lubrol-microdomains" contain the machinery for canalicular bile formation and may be the starting place for canalicular lipid secretion.

Endoscopic retrograde cholangiopancreaticography prior to explorative laparotomy avoids unnecessary surgery in patients suspected for biliary atresia.

BACKGROUND/AIMS: Timely diagnosis of biliary atresia (BA) requires key investigations that are less invasive but as accurate as possible. Non-invasive imaging preselects patients before explorative laparotomy is performed. The purpose of this prospective study was to evaluate the accuracy of endoscopic retrograde cholangiopancreaticography (ERCP) in these patients and to discuss its relevance to future diagnostic guidelines in neonatal jaundice. METHODS: Over a 7-year period, ERCP was routinely performed in cholestatic patients less than 6 months of age suspected for an extrahepatic origin of cholestasis, most likely BA. Endoscopic diagnosis was correlated with intraoperative findings. RESULTS: In 140 consecutive patients (mean age: 60 days; weight: 4 kg), ERCP excluded BA in 34 (25%) but failed in 18 newborns (13%) for technical reasons. The average procedure time was 23 min, and no severe complications occurred. Explorative laparotomy was performed in 106 patients and revealed BA in 80 cases. In this series, the sensitivity of ERCP for diagnosing biliary atresia was 92% and specificity was 73%. CONCLUSIONS: In preselected patients, ERCP is not an alternative to non-invasive imaging, but it avoids unnecessary surgical procedures in almost 25% of the cases. Hence, ERCP is recommended prior to explorative laparotomy in all patients suspected for BA.

5,5- and 5,6-Membered Spirocyclic Indolinone Hit-Finding Libraries.

The production of two libraries based on spirocyclic indolinones is described. These libraries were selected from numerous spirocyclic indolinone scaffolds with a library evaluation procedure used routinely at Novartis, based on computed physicochemical properties and measured properties of prototype compounds. The library production yielded 176 and 428 compounds that could be isolated in sufficient amounts and purities based on two closely related scaffolds. The novelty and diversity analysis of these libraries shows their complementarity to the chemical space covered by the structures of the PubChem database.

Increased susceptibility for intrahepatic cholestasis of pregnancy and contraceptive-induced cholestasis in carriers of the 1331T>C polymorphism in the bile salt export pump.

AIM: To study the association of three common ABCB11 and ABCC2 polymorphisms (ABCB11: 1331T>C --> V444A; ABCC2: 3563T>A --> V1188E and 4544G>A --> C1515Y) with intrahepatic cholestasis of pregnancy (ICP) and contraceptive-induced cholestasis (CIC). METHODS: ABCB11 and ABCC2 genotyping data were available from four CIC patients and from 42 and 33 ICP patients, respectively. Allele-frequencies of the studied polymorphisms were compared with those in healthy pregnant controls and Caucasian individuals. Furthermore, serum bile acid levels were correlated with the presence or absence of the 1331 C allele. RESULTS: The ABCB11 1331T>C polymorphism was significantly more frequent in cholestatic patients than in pregnant controls: C allele 76.2% (CI, 58.0-94.4) vs 51.3% (CI 35.8-66.7), respectively (P = 0.0007); and CC allele 57.1% (CI 36.0-78.3) vs 20% (CI 7.6-32.4), respectively (P = 0.0065). All four CIC patients were homozygous carriers of the C allele. In contrast, none of the studied ABCC2 polymorphism was overrepresented in ICP or CIC patients. Higher serum bile acid levels were found in carriers of the 1331CC genotype compared to carriers of the TT genotype. CONCLUSION: Our data support a role for the ABCB11 1331T>C polymorphism as a susceptibility factor for the development of estrogen-induced cholestasis, whereas no such association was found for ABCC2. Serum bile acid and gamma-glutamyl transferase levels might help to distinguish ABCB4- and ABCB11-related forms of ICP and CIC.

IgG4-related sclerosing cholangitis mimicking hilar cholangiocarcinoma (Klatskin tumor): a case report of a challenging disease and review of the literature.

BACKGROUND: Even though IgG4-related disease has gained increased attention worldwide, the diagnosis remains challenging. IgG4-related sclerosing cholangitis (IgG4-SC) is not well described in the western hemisphere and may mimic cholangiocarcinoma (CC), especially when occurring without other symptoms such as, e.g. concurrent pancreatitis or retroperitoneal fibrosis. We present a case to add further information to the diagnosis and treatment of this challenging disease. CASE REPORT: A 60-year-old male patient presented with painless jaundice. Prior medical history showed diabetes mellitus type I, high blood pressure, and deep vein thrombosis. Diagnostic investigations were strongly suspicious of a Klatskin tumor, although biopsies were inconclusive. The tumor marker Carbohydrate Antigen 19-9 (CA 19-9) was elevated. Prior to the recommended surgery, the patient had two second opinions in two different university hospitals, both arguing for surgery as well. The patient received hilar resection with right hemihepatectomy. During the postoperative course, some major complications occurred, i.e. recurrent pleural effusion, abscess in the liver resection area, sepsis, ileus, and restricted liver metabolism. Treatment with prednisolone did not show any improvement. Approximately 3 months after surgery, the patient died in consequence of acute respiratory failure. Histology showed no signs of CC, but IgG4-SC could be diagnosed. CONCLUSION: In the case of preoperative signs of CC, differential diagnosis of IgG4-SC needs to be considered, in particular, in cases with missing histologic proof of malignant disease.