GPCR kinase knockout cells reveal the impact of individual GRKs on arrestin binding and GPCR regulation.

G protein-coupled receptors (GPCRs) activate G proteins and undergo a complex regulation by interaction with GPCR kinases (GRKs) and the formation of receptor-arrestin complexes. However, the impact of individual GRKs on arrestin binding is not clear. We report the creation of eleven combinatorial HEK293 knockout cell clones lacking GRK2/3/5/6, including single, double, triple and the quadruple GRK knockout. Analysis of ß-arrestin1/2 interactions for twelve GPCRs in our GRK knockout cells enables the differentiation of two main receptor subsets: GRK2/3-regulated and GRK2/3/5/6-regulated receptors. Furthermore, we identify GPCRs that interact with ß-arrestins via the overexpression of specific GRKs even in the absence of agonists. Finally, using GRK knockout cells, PKC inhibitors and ß-arrestin mutants, we present evidence for differential receptor-ß-arrestin1/2 complex configurations mediated by selective engagement of kinases. We anticipate our GRK knockout platform to facilitate the elucidation of previously unappreciated details of GRK-specific GPCR regulation and ß-arrestin complex formation.

Phosphorus Analogues of [Ni(bpy)2]: Synthesis and Application in Carbon-Halogen Bond Activation.

The neutral, homoleptic pyridylphosphininenickel(0) complex [Ni(2-Py-4,6-Ph2-PC5H2)2] (1) has been obtained by reaction of the formal Ni(0) source [(IPr)Ni(H2C═CHSiMe3)2] with 2 equiv of 2-(2'-pyridyl)-4,6-diphenylphosphinine (L). Compound 1 can be oxidized both electrochemically and through the use of ferrocenium salts, to afford the corresponding Ni(I) complexes [1]BF4, [1(THF)]PF6, and [12](BArF4)2. The structures of these salts reveal an interesting dependence on the nature of the anion. While [1]BF4 and [1(THF)]PF6 show trigonal-bipyramidal coordination of Ni in the solid state, [12](BArF4)2 exists as a dinuclear Ni(I) complex and possesses a bridging phosphinine moiety in a rare µ2 mode. Reactions of 1 with halobenzenes highlight the noninnocent behavior of the aromatic phosphinine ligand, leading to the formation of oxidized Ni complexes but not to classical oxidative addition products. The reaction of 1 with bromobenzene affords the λ5 phosphinine 2 and the bipyramidal Ni(I) complex [1]Br, whereas a more unconventional oxidation product 3 is formed from the reaction of 1 and iodobenzene.

DNA methylation in the 5-HTT regulatory region is associated with CO2-induced fear in panic disorder patients.

A polymorphism in the gene encoding the serotonin (5-HT) transporter (5-HTT) has been shown to moderate the response to CO2 inhalation, an experimental model for panic attacks (PAs). Recurrent, unpredictable PAs represent, together with anticipatory anxiety of recurring attacks, the core feature of panic disorder (PD) and significantly interfere with patients' daily life. In addition to genetic components, accumulating evidence suggests that epigenetic mechanisms, which regulate gene expression by modifying chromatin structure, also play a fundamental role in the etiology of mental disorders. However, in PD, epigenetic mechanisms have barely been examined to date. In the present study, we investigated the relationship between methylation at the regulatory region of the gene encoding the 5-HTT and the reactivity to a 35% CO2 inhalation in PD patients. We focused on four specific CpG sites and found a significant association between the methylation level of one of these CpG sites and the fear response. This suggests that the emotional response to CO2 inhalation might be moderated by an epigenetic mechanism, and underlines the implication of the 5-HT system in PAs. Future studies are needed to further investigate epigenetic alterations in PD and their functional consequences. These insights can increase our understanding of the underlying pathophysiology and support the development of new treatment strategies.

Amino acid ferrocene conjugates using sulfonamide linkages.

This report presents the synthesis and characterization of mono- and bis(amino acid ester) ferrocene complexes generated using a sulfonamide linking strategy as an alternative to the more heavily explored amide linking strategy. These compounds were investigated to test their ability to form hydrogen bonding interactions both in the solid state and in solution, and were compared to the previously observed intramolecular interstrand crosslinking seen in amide-linked ferrocene constructs. Synthesized compounds also included controls that do not exhibit sulfonamide N-H bonds and thus cannot engage in hydrogen bonding. In the solid state, we observe both S=O⋯H-N and C=O⋯H-N intermolecular interactions, but we do not observe any intramolecular interstrand hydrogen bonding. In the solution phase, we also do not see any intramolecular hydrogen bonding interactions in these compounds as measured by titration of d6-DMSO as a competitive hydrogen bonding reagent. We also collected CD spectra on these compounds, which revealed that the chiral peptides can induce dichroism in the dd transition of the ferrocene units. Our results indicate that the peptide-ferrocene linking group governs whether intermolecular hydrogen bonding interactions can occur between the amino acids adjacent to the cyclopentadienyl groups.

MAOA gene hypomethylation in panic disorder-reversibility of an epigenetic risk pattern by psychotherapy.

Epigenetic signatures such as methylation of the monoamine oxidase A (MAOA) gene have been found to be altered in panic disorder (PD). Hypothesizing temporal plasticity of epigenetic processes as a mechanism of successful fear extinction, the present psychotherapy-epigenetic study for we believe the first time investigated MAOA methylation changes during the course of exposure-based cognitive behavioral therapy (CBT) in PD. MAOA methylation was compared between N=28 female Caucasian PD patients (discovery sample) and N=28 age- and sex-matched healthy controls via direct sequencing of sodium bisulfite-treated DNA extracted from blood cells. MAOA methylation was furthermore analyzed at baseline (T0) and after a 6-week CBT (T1) in the discovery sample parallelized by a waiting time in healthy controls, as well as in an independent sample of female PD patients (N=20). Patients exhibited lower MAOA methylation than healthy controls (P<0.001), and baseline PD severity correlated negatively with MAOA methylation (P=0.01). In the discovery sample, MAOA methylation increased up to the level of healthy controls along with CBT response (number of panic attacks; T0-T1: +3.37±2.17%), while non-responders further decreased in methylation (-2.00±1.28%; P=0.001). In the replication sample, increases in MAOA methylation correlated with agoraphobic symptom reduction after CBT (P=0.02-0.03). The present results support previous evidence for MAOA hypomethylation as a PD risk marker and suggest reversibility of MAOA hypomethylation as a potential epigenetic correlate of response to CBT. The emerging notion of epigenetic signatures as a mechanism of action of psychotherapeutic interventions may promote epigenetic patterns as biomarkers of lasting extinction effects.

Systematic review and meta-analysis of hemodynamic-directed feedback during cardiopulmonary resuscitation in cardiac arrest.

BACKGROUND/OBJECTIVE: Physiologic monitoring of resuscitative efforts during cardiac arrest is gaining in importance, as it provides a real-time window into the cellular physiology of patients. The aim of this review is to assess the quality of evidence surrounding the use of physiologic monitoring to guide cardiopulmonary resuscitation (CPR), and to examine whether the evidence demonstrates an improvement in patient outcome when comparing hemodynamic-directed CPR versus standard CPR. METHODS: Studies were obtained through a search of the PubMed, Embase and Cochrane databases. Peer-reviewed randomized trials, case-control studies, systematic reviews, and cohort studies that titrated CPR to physiologic measures, compared results to standard CPR, and examined patient outcome were included. RESULTS: Six studies met inclusion criteria, with all studies conducted in animal populations. Four studies examined the effects of hemodynamic-directed CPR on survival, with 35/37 (94.6%) animals surviving in the hemodynamic-directed CPR groups and 12/35 (34.3%) surviving in the control groups (p<0.001). Two studies examined the effects of hemodynamic-directed CPR on ROSC, with 22/30 (73.3%) achieving ROSC in the hemodynamic-directed CPR group and 19/30 (63.3%) achieving ROSC in the control group (p=0.344). DISCUSSION/CONCLUSION: These results suggest a trend in survival from hemodynamic-directed CPR over standard CPR, however the small sample size and lack of human data make these results of limited value. Future human studies examining hemodynamic-directed CPR versus current CPR standards are needed to enhance our understanding of how to effectively use physiologic measures to improve resuscitation efforts and ultimately incorporate concrete targets into international resuscitation guidelines.

Clonal B cells in Waldenström's macroglobulinemia exhibit functional features of chronic active B-cell receptor signaling.

Waldenström's macroglobulinemia (WM) is a B-cell non-Hodgkin's lymphoma (B-NHL) characterized by immunoglobulin M (IgM) monoclonal gammopathy and the medullary expansion of clonal lymphoplasmacytic cells. Neoplastic transformation has been partially attributed to hyperactive MYD88 signaling, secondary to the MYD88 L265P mutation, occurring in the majority of WM patients. Nevertheless, the presence of chronic active B-cell receptor (BCR) signaling, a feature of multiple IgM+ B-NHL, remains a subject of speculation in WM. Here, we interrogated the BCR signaling capacity of primary WM cells by utilizing multiparametric phosphoflow cytometry and found heightened basal phosphorylation of BCR-related signaling proteins, and augmented phosphoresponses on surface IgM (sIgM) crosslinking, compared with normal B cells. In support of those findings we observed high sIgM expression and loss of phosphatase activity in WM cells, which could both lead to signaling potentiation in clonal cells. Finally, led by the high-signaling heterogeneity among WM samples, we generated patient-specific phosphosignatures, which subclassified patients into a 'high' and a 'healthy-like' signaling group, with the second corresponding to patients with a more indolent clinical phenotype. These findings support the presence of chronic active BCR signaling in WM while providing a link between differential BCR signaling utilization and distinct clinical WM subgroups.

[Surgical treatment of ulnar collateral ligament injuries of the thumb metacarpophalangeal joint]. / Operative Behandlung der ulnaren Seitenbandläsion am Daumengrundgelenk.

OBJECTIVE: Reconstruction of the ruptured ulnar collateral ligament of the metacarpophalangeal (MP) joint of the thumb. INDICATIONS: Ruptured ulnar collateral ligament of the thumb MP joint with instability: joint opening of more than 30° in flexion and more than 20° in extension, Stener lesion, displaced avulsion fractures. CONTRAINDICATIONS: Abrasions, wound-healing disturbance, skin disease, osteoarthritis. SURGICAL TECHNIQUE: Curved skin incision dorsoulnar above the thumb MP joint. Protection of the branches of the superficial radial nerve. Incision of the adductor aponeurosis. Exposing the ulnar collateral ligament; opening and examination of the joint. Depending on the injury, primary suture repair, transosseous suture, repair with a bone anchor, osteosynthesis with K-wires or small screws in avulsion fracture, ligament reconstruction in chronic instability or older injury. POSTOPERATIVE TREATMENT: Cast splint of the MP joint until swelling subsides; cast immobilization for 6 weeks; range-of-motion exercises, avoiding forced radial deviation of the MP joint for 3 months. RESULTS: Complete joint stability 3 months postoperatively in all 34 patients with rupture of the ulnar collateral ligament.

Attachment style and oxytocin receptor gene variation interact in influencing social anxiety.

OBJECTIVES: Social anxiety has been suggested to be promoted by an insecure attachment style. Oxytocin is discussed as a mediator of trust and social bonding as well as a modulator of social anxiety. Applying a gene-environment (G × E) interaction approach, in the present pilot study the main and interactive effects of attachment styles and oxytocin receptor (OXTR) gene variation were probed in a combined risk factor model of social anxiety in healthy probands. METHODS: Participants (N = 388; 219 females, 169 males; age 24.7 ± 4.7 years) were assessed for anxiety in social situations (Social Phobia and Anxiety Inventory) depending on attachment style (Adult Attachment Scale, AAS) and OXTR rs53576 A/G genotype. RESULTS: A less secure attachment style was significantly associated with higher social anxiety. This association was partly modulated by OXTR genotype, with a stronger negative influence of a less secure attachment style on social anxiety in A allele carriers as compared to GG homozygotes. CONCLUSIONS: The present pilot data point to a strong association of less secure attachment and social anxiety as well as to a gene-environment interaction effect of OXTR rs53576 genotype and attachment style on social anxiety possibly constituting a targetable combined risk marker of social anxiety disorder.

Hydrazine-mediated strongly coupled Re(CO)3 dimers.

Dimeric metal complexes can often exhibit coupling interactions via bridging ligands. In this report, we present two Re(CO)3 dimers, where the metals are linked via a bis(pyca) hydrazine (pyca = pyridine-2-carbaldehyde imine) Schiff base ligand. For the dimeric compounds 4 and 5, we observe strong coupling across the dimer as measured by cyclic voltammetry: ∼480 mV separations between the first and the second reduction waves that correspond to comproportionation constants close to 1.5 × 10(8). Evidence for a mixed valence state upon one electron reduction was also observed by spectroelectrochemistry in which a clear inter-valence charge-transfer (IVCT) band was observed in [4]- and [5]-complexes. The electronic structures of all target compounds were probed by DFT and TDDFT computational methods. DFT calculations indicate that reduction takes place at the diimine units, and that the observed coupling is a ligand-based phenomenon, rather than one that involves metal-based orbitals.

Induction of E/Z isomerization in a pendant metal-bound azobenzene: a synthetic, spectroscopic and theoretical study.

Re(CO)3 conjugates 1 and 2 that incorporate azobenzenes can be readily generated via one-pot reactions using Schiff base reaction forming conditions. Excitation of the MLCT bands in 1 and 2 results in isomerization of the azobenzene moiety, and this process has been investigated via time-resolved photophysics and TDDFT calculations.

Lithium-induced Clock Gene Expression in Lymphoblastoid Cells of Bipolar Affective Patients.

INTRODUCTION: Disturbances of circadian rhythms occur in all episodes of bipolar disorder (BD). Lithium, as gold-standard in the maintenance treatment of BD, is known to influence circadian processes. METHODS: In a pilot study lymphoblastoid cell lines (LCLs) were generated from 8 BD patients and 6 healthy controls. The LCLs were treated with lithiumchloride (LiCl) for 3 weeks. Cell cycles were then synchronized and expressional analysis by quantitative Real Time PCR was done. RESULTS: BD and controls differed in the period length regarding DBP (albumin D-box binding protein) expression and DBP expression was also influenced by lithium treatment. Furthermore, baseline DBP expression was significantly different between non-treated BD and healthy controls. None of the other analyzed circadian genes showed to be influenced by chronic lithium treatment or to be differentially regulated due to the diagnosis. DISCUSSION: We here show that chronic lithium treatment of LCLs leads to decreased expression of the clock gene DBP, rendering DBP a lithium-regulated gene. We could confirm the role of the circadian clock as well in lithium mode of action as in the pathomechanisms of BD although future studies with a greater number of participants and cell lines are needed.

Ultrafast dynamics of a new class of highly fluorescent boron difluoride dyes.

The first ultrafast study of the dimeric fluorescent BF2 dye BOPHY is presented. When compared to a structurally related BODIPY dye, similar photophysical dynamics are observed, including an intermediate kinetic component present in both dye types.

Charge-induced distortion and stabilization of surface transfer doped porphyrin films.

The interaction between zinc-tetraphenylporphyrin (ZnTPP) and fullerenes (C60 and C60F48) are studied using ultraviolet photoelectron spectroscopy (UPS) and scanning tunneling microscopy (STM). Low temperature STM reveals highly ordered ZnTPP monolayers on Au(111). In contrast to C60, a submonolayer coverage of C60F48 results in long-range disorder of the underlying single ZnTPP layer and distortion of individual ZnTPP molecules. This is induced by substantial charge transfer at the organic-organic interface, revealed by the interface energetics from UPS. However, a second layer of ZnTPP prevents C60F48 guests from breaking the self-assembled porphyrin template. This finding is important for understanding the growth behaviour of "bottom-up" functional nanostructures involving strong donor-acceptor heterojunctions in molecular electronics.

Selectivity mechanism of the mechanosensitive channel MscS revealed by probing channel subconducting states.

The mechanosensitive channel of small conductance (MscS) has been characterized at both functional and structural levels and has an integral role in the protection of bacterial cells against hypoosmotic shock. Here we investigate the role that the cytoplasmic domain has in MscS channel function by recording wild-type and mutant MscS single-channel activity in liposome patches. We report that MscS preferentially resides in subconducting states at hyperpolarising potentials when Ca(2+) and Ba(2+) ions are the major permeant cations. In addition, our results indicate that charged residues proximal to the seven vestibular portals and their electrostatic interactions with permeating cations determine selectivity and regulate the conductance of MscS and potentially other channels belonging to the MscS subfamily. Furthermore, our findings suggest a role for mechanosensitive channels in bacterial calcium regulation, indicative of functions other than protection against osmolarity changes that these channels possibly fulfil in bacteria.

[Pediatrics. Sharing of the medical letter with the patient]. / Pédiatrie. 5. Le partage de l'information médicate en 2012.

Some paediatrics centres routinely send the medical consultation letter not only to the primary or referring physician but to the patient and his/her family as well. This way of sharing medical information is appreciated not only by the patients themselves but also by a majority of physicians.

Anti-tumor effects of peptide analogs targeting neuropeptide hormone receptors on mouse pheochromocytoma cells.

Pheochromocytoma is a rare but potentially lethal chromaffin cell tumor with currently no effective treatment. Peptide hormone receptors are frequently overexpressed on endocrine tumor cells and can be specifically targeted by various anti-tumor peptide analogs. The present study carried out on mouse pheochromocytoma cells (MPCs) and a more aggressive mouse tumor tissue-derived (MTT) cell line revealed that these cells are characterized by pronounced expression of the somatostatin receptor 2 (sst2), growth hormone-releasing hormone (GHRH) receptor and the luteinizing hormone-releasing hormone (LHRH) receptor. We further demonstrated significant anti-tumor effects mediated by cytotoxic somatostatin analogs, AN-162 and AN-238, by LHRH antagonist, Cetrorelix, by the cytotoxic LHRH analog, AN-152, and by recently developed GHRH antagonist, MIA-602, on MPC and for AN-152 and MIA-602 on MTT cells. Studies of novel anti-tumor compounds on these mouse cell lines serve as an important basis for mouse models of metastatic pheochromocytoma, which we are currently establishing.

Quality of life in patients with muscle invasive and non-muscle invasive bladder cancer.

PURPOSE: Compared to the literature on other malignancies, data on quality of life (QoL) in bladder cancer are sparse. This study sought answers to the following questions: In what QoL domains do patients with bladder cancer differ from the general population? Do patients with radical cystectomy differ in QoL compared to those who received conservative treatment? Do patients with neobladder generally have better QoL compared to patients with other diversion methods? METHODS: At the beginning of inpatient rehabilitation, N = 823 patients with bladder cancer were assessed. Data of a representative community sample (N = 2037) were used for comparison. The questionnaire EORTC QLQ-C30 was used to measure QoL. Multivariate linear regression models were computed to investigate differences between groups. RESULTS: Patients with both non-muscle invasive and muscle invasive bladder cancer reported significantly more problems and worse functioning than the general population. Radiotherapy is associated with clinically relevant more pain, dyspnoea, constipation, appetite loss and decreased social functioning while chemotherapy is associated more with dyspnoea. Cystectomy patients reported more fatigue, appetite loss and decreased role functioning. Male patients ≥70 years with conduit experienced more sleep and emotional problems. These effects of urinary diversion were not observed in women and younger patients. CONCLUSIONS: Patients with bladder cancer experience various QoL concerns at the beginning of inpatient rehabilitation. These problems can partly be explained by the type of treatment the patients receive. Type of urinary diversion is relevant for QoL in subgroups of patients.

[Therapy-resistant aggressive cystic lesion of the mandible]. / Therapieresistente aggressive zystische Läsion des Unterkiefers.

Several extensive surgical interventions of a cystic lesion in the left mandible were followed by recurrences. The lesion extended from the primary mandibular region into the area of pterygopalatine fossa finally infiltrating the orbital region and the skull base. Histological results could never demonstrate a malignancy with certainty. Due to the patient's poor general condition, the refusal for further surgical inventions and due to the malignoma-like growth pattern radiation treatment was performed. However, this had no effect on tumor progression.

Highly transparent Bi2MoO6- and Bi2WO6-polymer nanocomposites.

A combined method of precipitation, phase transfer into organic solvent, solvothermal treatment and subsequent in situ polymerization was used to integrate nanocrystalline Bi2MoO6- and Bi2WO6-particles into a polymer matrix of poly-laurylacrylate. The presented method offers a new and gentle way to produce highly transparent bulk nanocomposites containing evenly distributed Bi2MoO6- and Bi2WO6-nanoparticles. Characterization results of DLS-, XRD-, REM- and TEM-measurements are presented as well as solid state UV/VIS-measurements of the particles. The transparent nanocomposites were characterized using UV/VIS-spectroscopy and ellipsometry. All composites show a good transmission in the range from 800-400 nm. The particle content of the nanocomposites was measured with TG-measurements.