Regulatory modules mediating the complex neural expression patterns of the homeobrain gene during Drosophila brain development.

BACKGROUND: The homeobox gene homeobrain (hbn) is located in the 57B region together with two other homeobox genes, Drosophila Retinal homeobox (DRx) and orthopedia (otp). All three genes encode transcription factors with important functions in brain development. Hbn mutants are embryonic lethal and characterized by a reduction in the anterior protocerebrum, including the mushroom bodies, and a loss of the supraoesophageal brain commissure. RESULTS: In this study we conducted a detailed expression analysis of Hbn in later developmental stages. In the larval brain, Hbn is expressed in all type II lineages and the optic lobes, including the medulla and lobula plug. The gene is expressed in the cortex of the medulla and the lobula rim in the adult brain. We generated a new hbnKOGal4 enhancer trap strain by reintegrating Gal4 in the hbn locus through gene targeting, which reflects the complete hbn expression during development. Eight different enhancer-Gal4 strains covering 12 kb upstream of hbn, the two large introns and 5 kb downstream of the gene, were established and hbn expression was investigated. We characterized several enhancers that drive expression in specific areas of the brain throughout development, from embryo to the adulthood. Finally, we generated deletions of four of these enhancer regions through gene targeting and analysed their effects on the expression and function of hbn. CONCLUSION: The complex expression of Hbn in the developing brain is regulated by several specific enhancers within the hbn locus. Each enhancer fragment drives hbn expression in several specific cell lineages, and with largely overlapping patterns, suggesting the presence of shadow enhancers and enhancer redundancy. Specific enhancer deletion strains generated by gene targeting display developmental defects in the brain. This analysis opens an avenue for a deeper analysis of hbn regulatory elements in the future.

Functional analysis of enhancer elements regulating the expression of the Drosophila homeodomain transcription factor DRx by gene targeting.

BACKGROUND: The Drosophila brain is an ideal model system to study stem cells, here called neuroblasts, and the generation of neural lineages. Many transcriptional activators are involved in formation of the brain during the development of Drosophila melanogaster. The transcription factor Drosophila Retinal homeobox (DRx), a member of the 57B homeobox gene cluster, is also one of these factors for brain development. RESULTS: In this study a detailed expression analysis of DRx in different developmental stages was conducted. We show that DRx is expressed in the embryonic brain in the protocerebrum, in the larval brain in the DM and DL lineages, the medulla and the lobula complex and in the central complex of the adult brain. We generated a DRx enhancer trap strain by gene targeting and reintegration of Gal4, which mimics the endogenous expression of DRx. With the help of eight existing enhancer-Gal4 strains and one made by our group, we mapped various enhancers necessary for the expression of DRx during all stages of brain development from the embryo to the adult. We made an analysis of some larger enhancer regions by gene targeting. Deletion of three of these enhancers showing the most prominent expression patterns in the brain resulted in specific temporal and spatial loss of DRx expression in defined brain structures. CONCLUSION: Our data show that DRx is expressed in specific neuroblasts and defined neural lineages and suggest that DRx is another important factor for Drosophila brain development.

The Drosophila homeodomain transcription factor Homeobrain is involved in the formation of the embryonic protocerebrum and the supraesophageal brain commissure.

During the embryonic development of Drosophila melanogaster many transcriptional activators are involved in the formation of the embryonic brain. In our study we show that the transcription factor Homeobrain (Hbn), a member of the 57B homeobox gene cluster, is an additional factor involved in the formation of the embryonic Drosophila brain. Using a Hbn antibody and specific cell type markers a detailed expression analysis during embryonic brain development was conducted. We show that Hbn is expressed in several regions in the protocerebrum, including fibre tract founder cells closely associated with the supraesophageal brain commissure and also in the mushroom bodies. During the formation of the supraesophageal commissure, Hbn and FasII-positive founder cells build an interhemispheric bridge priming the commissure and thereby linking both brain hemispheres. The Hbn expression is restricted to neural but not glial cells in the embryonic brain. In a mutagenesis screen we generated two mutant hbn alleles that both show embryonic lethality. The phenotype of the hbn mutant alleles is characterized by a reduction of the protocerebrum, a loss of the supraesophageal commissure and mushroom body progenitors and also by a dislocation of the optic lobes. Extensive apoptosis correlates with the impaired formation of the embryonic protocerebrum and the supraesophageal commissure. Our results show that Hbn is another important factor for embryonic brain development in Drosophila melanogaster.

The homeodomain transcription factor Orthopedia is involved in development of the Drosophila hindgut.

BACKGROUND: The Drosophila hindgut is commonly used model for studying various aspects of organogenesis like primordium establishment, further specification, patterning, and morphogenesis. During embryonic development of Drosophila, many transcriptional activators are involved in the formation of the hindgut. The transcription factor Orthopedia (Otp), a member of the 57B homeobox gene cluster, is expressed in the hindgut and nervous system of developing Drosophila embryos, but due to the lack of mutants no functional analysis has been conducted yet. RESULTS: We show that two different otp transcripts, a hindgut-specific and a nervous system-specific form, are present in the Drosophila embryo. Using an Otp antibody, a detailed expression analysis during hindgut development was carried out. Otp was not only expressed in the embryonic hindgut, but also in the larval and adult hindgut. To analyse the function of otp, we generated the mutant otp allele otpGT by ends-out gene targeting. In addition, we isolated two EMS-induced otp alleles in a genetic screen for mutants of the 57B region. All three otp alleles showed embryonic lethality with a severe hindgut phenotype. Anal pads were reduced and the large intestine was completely missing. This phenotype is due to apoptosis in the hindgut primordium and the developing hindgut. CONCLUSION: Our data suggest that Otp is another important factor for hindgut development of Drosophila. As a downstream factor of byn Otp is most likely present only in differentiated hindgut cells during all stages of development rather than in stem cells.

Retinal homeobox promotes cell growth, proliferation and survival of mushroom body neuroblasts in the Drosophila brain.

The Drosophila mushroom bodies, centers of olfactory learning and memory in the fly 'forebrain', develop from a set of neural stem cells (neuroblasts) that generate a large number of Kenyon cells (KCs) during sustained cell divisions from embryonic to late pupal stage. We show that retinal homeobox (rx), encoding for an evolutionarily conserved transcription factor, is required for proper development of the mushroom bodies. Throughout development rx is expressed in mushroom body neuroblasts (MBNBs), their ganglion mother cells (MB-GMCs) and young KCs. In the absence of rx function, MBNBs form correctly but exhibit a reduction in cell size and mitotic activity, whereas overexpression of rx increases growth of MBNBs. These data suggest that Rx is involved in the control of MBNB growth and proliferation. Rx also promotes cell cycling of MB-GMCs. Moreover, we show that Rx is important for the survival of MBNBs and Kenyon cells which undergo premature cell death in the absence of rx function. Simultaneous blocking of cell death restores the normal set of MBNBs and part of the KCs, demonstrating that both, impaired proliferation and premature cell death (of MBNBs and KCs) account for the observed defects in mushroom body development. We then show that Rx controls proliferation within the MBNB clones independently of Tailless (Tll) and Prospero (Pros), and does not regulate the expression of other key regulators of MB development, Eyeless (Ey) and Dachshund (Dac). Our data support that the role of Rx in forebrain development is conserved between vertebrates and fly.

Restructuring of an Ir(210) electrode surface by potential cycling.

This study addresses the electrochemical surface faceting and restructuring of Ir(210) single crystal electrodes. Cyclic voltammetry measurements and in situ scanning tunnelling microscopy are used to probe structural changes and variations in the electrochemical behaviour after potential cycling of Ir(210) in 0.1 M H2SO4. Faceted structures are obtained electrochemically as a function of time by cycling at a scanrate of 1 V·s(-1) between -0.28 and 0.70 V vs SCE, i.e., between the onset of hydrogen evolution and the surface oxidation regime. The electrochemical behaviour in sulfuric acid solution is compared with that of thermally faceted Ir(210), which shows a sharp characteristic voltammetric peak for (311) facets. Structures similar to thermally-induced faceted Ir(210) are obtained electrochemically, which typically correspond to polyoriented facets at nano-pyramids. These structures grow anisotropically in a preferred direction and reach a height of about 5 nm after 4 h of cycling. The structural changes are reflected in variations of the electrocatalytic activity towards carbon monoxide adlayer oxidation.

Roughening and long-range nanopatterning of Au(111) through potential cycling in aqueous acidic media.

Electrochemical treatment of Au(111) in aqueous H2SO4 solution by repetitive application of oxide formation-reduction cycles (OFRC) generates nanopatterned surfaces with long-range order. The pattern development depends on the lower and upper potential limits (EL, EU), the number (n) of OFRCs, and the potential scan rate (s). Surface patterning of Au(111) initially (n = 1-2) generates small islands and holes that are one atomic step in height. As n increases to 5, the number of islands decreases and the holes become larger; after n = 10 OFRCs, the islands become inexistent and large, randomly distributed holes are observed. Increase of OFRCs to n = 20 generates surface structures that reside within three atomic layers and resemble phase separation through a spinodal decomposition mechanism. As the number of OFRCs rises to n = 50, a network of interconnected islands and holes emerges; the islands and holes are two-three atomic steps in height, and are located within topmost five monolayers. Further increase of the number of OFRCs to n = 100 creates a network of interconnected trigonal pyramids that are pointed in the same direction. The size of the pyramids depends on the electrolyte composition and the number of OFRCs. In the case of n = 100, the pyramids are 12-25 nm in base length and 0.4-1.6 nm in height in 0.1 M aqueous H2SO4, and 20-50 nm in base length and 0.8-1.6 nm in height in 0.1 M aqueous HNO3. The number of OFRCs and scan rate play an important role in patterning of Au(111), and complete nanopattern development requires a large number of OFRCs and low scan rates.

The interfaces of Au(111) and Au(100) in a hexaalkyl-substituted guanidinium ionic liquid: an electrochemical and in situ STM study.

Five hexaalkylguanidinium-based ionic liquids have been synthesised, and based on their cyclic voltammograms the most suited one, N,N-dibutyl-N',N'-diethyl-N'',N''-dimethylguanidinium bis(trifluoromethylsulfonyl)imide, has been chosen for electrochemical studies. The surface interaction of this room-temperature ionic liquid with single crystalline gold surfaces (Au(100) and Au(111)) has been investigated using cyclic voltammetry, impedance spectroscopy and in situ scanning tunnelling microscopy (STM). The interfacial capacitance was found to be very low; STM measurements revealed the hex-reconstruction and herringbone reconstruction for Au(100) and for Au(111), respectively, at negative potentials; that is, at these potentials no hints for ad-structures of the cation could be found.

Enzyme immobilisation on self-organised nanopatterned electrode surfaces.

A new method is described for immobilisation of enzymes on polymer-coated Pt islands. These islands are deposited on top of a SAM-covered Au(111) electrode by a combination of electroless and electrochemical deposition, which allows for a variation of island size and distance between the islands. Here we describe the immobilisation of pyranose-2-oxidase (P2Ox) and the catalytic response to D-glucose on such a nanopatterned surface, which provides optimum access to the active centres of the enzyme.

Metallization of ultra-thin, non-thiol SAMs with flat-lying molecular units: Pd on 1, 4-dicyanobenzene.

Self-assembled monolayers of 1,4-dicyanobenzene on Au(111) electrodes are studied by cyclic voltammetry, in-situ STM and ex-situ XPS. High-resolution STM images reveal a long-range order of propeller-like assemblies each of which consists of three molecules, all lying flat on the gold substrate with the cyano groups oriented parallel to the metal surface. It is demonstrated that both functional groups can act as complexation sites for metal ions from solution. Surprisingly, such arrangements still allow the metal to be deposited on top of the molecules by electrochemical reduction despite the close vicinity to the Au surface. The latter is demonstrated by angle-resolved XPS which unequivocally shows that the metal indeed resides on top of the organic layer rather than underneath, despite the flat arrangement of the molecules.

Metallization of organic surfaces: Pd on thiazole.

Results from electrochemical studies, in situ STM, and ex situ angle-resolved XPS on self-assembled monolayers (SAMs) of thiazole on Au(111) are reported for the first time. Although STM seems to indicate a low density of molecules organized in small ordered domains, a quantitative chemical analysis of the sample surface by XPS clearly points toward the formation of a densely packed molecular layer. The stability of the thiazole SAM against reductive desorption is found to be very comparable with that for thiol-SAMs on gold. This results from the formation of Au-S bonds between the molecules and their support as evidenced by XPS, thereby rebuting speculations that the ring nitrogen is responsible for the attachment of such molecules to gold surfaces. Consequently, the N-atoms terminating the molecular layer are available as active sites for the complexation with Pd ions thereby allowing the deposition of Pd islands with monatomic height on top of the thiazole SAM. The importance of such studies for metal-molecule interconnections is briefly addressed.

Prognostic factors of liver injury in polytraumatic patients. Results from 895 severe abdominal trauma cases.

BACKGROUND AND AIMS: Prognosis of multiple injured patients is mainly limited by severe haemorrhage. Although mechanisms of altered immune response have been intensively investigated, little is known about the relevance of liver trauma as an independent predictive outcome factor in these patients. METHODS: 10,469 patients from the DGU Trauma Registry (1993-2005) were retrospectively analyzed. Primary admitted patients with an injury severity score > or = 16, without isolated head injury were included. Patients were analyzed according to the injury pattern as liver injury (Abbreviated Injury Scale--AIS abdomen < 3 and AIS liver 2-5; n = 321), non-liver abdominal trauma (AIS abdomen 2-5 or AIS liver < 3; n = 574) and control group without abdominal injuries (AIS abdomen or liver < 3; n = 9,574). RESULTS: Severe liver injury was associated with excessive demands for volume resuscitation and induced a significantly increased risk for sepsis and multi-organ failure (MOF) compared to both other groups (sepsis 19.9% vs. 11.0%; MOF 32.7% vs. 16.6%). Furthermore, deleterious outcome was more frequently associated with severe liver trauma (mortality 34.9%) compared to severe abdominal trauma (12.0%). CONCLUSION: Severe liver trauma is an independent predictor for severe haemorrhage with a substantially increased risk of sepsis, MOF and trauma-related death. While conservative treatment of patients with liver trauma but no haemorrhage is effective, patients with hemodynamic instability seem to be from a subgroup where contemporary treatment modalities are not yet sufficient.

Major trauma with multiple injuries in German children: a retrospective review.

BACKGROUND: The aim of this study was to evaluate the demographics, mechanisms, pattern, injury severity, and the outcome (ie, length of intensive care unit [ICU] stay, length of mechanical ventilation, total length of stay, mortality) in multiple-injured children based on a review from the German trauma registry study ("Traumaregister") of the German Society of Trauma Surgery (Deutsche Gesellschaft für Unfallchirurgie e.V.). METHODS: One hundred three German trauma centers took part in the German trauma registry study from January 1997 to December 2003. Five hundred seventeen children (aged 0-15 years) with multiple injuries and an Injury Severity Score of more than 15 in comparison to 11,025 adults were included. Sex, age, and mechanisms and pattern of injury were assessed. The mechanisms of trauma and the anatomical distribution of severe injury (Abbreviated Injury Scale of 3 or more) were analyzed. The Injury Severity Score, the Revised Trauma Score, and the Trauma Score Injury Severity Score were calculated to estimate the severity of injury and mortality. RESULTS: The predominant sex was male. Most cases were caused by traffic-related accidents. Head injuries were most common in children, and severe thoracic injuries increased with age. Mean length of ICU treatment, mechanical ventilation, and total length of stay were shorter in children than in adults. A total of 22.6% of the children aged 0 to 5 years died in the hospital in comparison with in-hospital mortality rate of 13.7% in the 6- to 10-, 20.3% in the 11- to 15-, and 17.0% in the 16- to 55-year-old patients. CONCLUSIONS: There were differences between multiple-injured children and adults concerning injury mechanisms and pattern of injuries. Adults needed a longer mechanical ventilation and a longer ICU therapy. Most deaths could be seen in the youngest patients aged 0 to 5 years.

Reduction in mortality of severely injured patients in Germany.

INTRODUCTION: The trauma registry of the German Society of Trauma Surgery is a multicentric prospective record of the treatment of severely injured patients. METHODS: The present study examines the effect of a quality management system on key processes and outcomes, in hospitals included in the trauma registry. The study is based on data of 11 013 severely injured patients (injury severity score = 16) who were treated in 105 hospitals between 1993 and 2005. A variety of parameters relating to early diagnosis and treatment were considered. Outcome quality was measured by a comparison between observed and calculated mortality (revised injury severity classification). RESULTS: During the 13 year long study period mortality could be significantly reduced from 22.8% to 18.7%. The time to initial radiological and ultrasound diagnosis was reduced, the use of computed tomography increased, the time until emergency operations in hemorrhagic shock was reduced, and damage limiting orthopedic interventions were performed more frequently. DISCUSSION: The German Trauma Registry records processes and treatment results in severely injured patients. This information is fed back to participating hospitals. The continuous data feedback is associated with a continuous improvement of process and outcome quality in the treatment of severely injured patients.

Potential-induced structure changes of oligopyridine adlayers on Au(111) electrodes.

The structure of a bisterpyridine-like oligopyridine (abbreviated as 2,4'-BTP) monolayer on Au(111), adsorbed from an acetone solution, was studied by in situ scanning tunneling microscopy and cyclic voltammetry in aqueous 0.1 M H2SO4. Short-range ordered adsorption with an average distance between the individual molecules of about 2 nm was observed only at electrode potentials positive of +0.4 V vs SCE, whereas at more negative potentials, no order could be found. With the help of Cu underpotential deposition, a potential-induced, fast, and fully reversible structure transition within the organic monolayer was identified at about +0.4 V vs SCE. At negative potentials the molecules apparently cluster together and consequently current-potential curves resemble those for a bare gold surface, whereas for E>+0.4 V vs SCE the molecules are spread over the entire surface in a hexagonal, close-packed fashion. This may have interesting consequences for switching between different template structures.

Pelvic Ring Fractures: Utility of Clinical Examination in Patients with Impaired Consciousness or Tracheal Intubation.

OBJECTIVE: To define the diagnostic accuracy of clinical examination in patients with impaired consciousness or endotracheal intubation to detect pelvic ring fractures and to identify those with severe bleeding. METHODS: Included in this prospective data collection with retrolective data analysis were a consecutive series of blunt trauma victims with either a Glasgow Coma Scale ≤ 13 or tracheal intubation. Clinical examination comprised testing for stability of the iliac wings. RESULTS: From 784 subjects (injury severity score 23.3 ± 17.4) 93 patients (11.9%) were found to have a pelvic ring fracture. Clinical instability of the pelvic ring was found in 42 patients. There was only one false positive. Fifty-two fractures could not be identified by clinical examination, including nine fractures (17%) that required surgical fracture stabilization (sensitivity of clinical examination 44.1%). Seventeen fractures (18.3%) were associated with a blood loss larger than 20% of circulating blood volume. Sixteen of those were identified by clinical instability of the pelvic ring (sensitivity 94.1%, specificity 97.0%, positive predictive value 38.1%, negative predictive value 99.9%). CONCLUSIONS: Clinical examination for stability of the pelvis in this selected group of patients missed a significant number of pelvic ring fractures including fractures that require surgical stabilization. The finding of a clinically unstable identifies most of the patients with the pelvic ring fracture being a major source of bleeding. A stable pelvis makes pelvic ring fracture as being the source of bleeding quite unlikely.