Development of spinning-disk solid sample delivery system for high-repetition rate x-ray free electron laser experiments.

X-ray free-electron lasers (XFELs) deliver intense x-ray pulses that destroy the sample in a single shot by a Coulomb explosion. Experiments using XFEL pulse trains or the new generation of high-repetition rate XFELs require rapid sample replacement beyond those provided by the systems now used at low repletion-rate XFELs. We describe the development and characterization of a system based on a spinning disk to continuously deliver a solid sample into an XFEL interaction point at very high speeds. We tested our system at the Linac Coherent Light Source and European XFEL hard x-ray nano-focus instruments, employing it to deliver a 25 µm copper foil sample, which can be used as a gain medium for stimulated x-ray emission for the proposed x-ray laser oscillator.

In-office outpatient orthognathic surgery: review of 254 cases where the patients were discharged the same day.

This study was performed to present a single operator's experience of in-office (outside of a hospital setting) outpatient orthognathic surgery over a period of 12 years. A total of 254 surgeries were performed during this period. Average procedure times were comparable with published results from studies of similar material. The mean operating time for bimaxillary surgery (n = 21) was 3 hours and 11 minutes. Regarding single-jaw procedures, the mean operating time for Le Fort I osteotomy (n = 115) was 2 hours and 14 minutes and for bilateral sagittal split osteotomy (n = 118) was 2 hours and 1 minute. All patients were discharged from the office the same day, except one patient who was transported to the hospital after surgery due to an anaesthetic complication. This patient was discharged from the hospital later the same day. In this setting, outpatient orthognathic surgery is both safe and practical when careful attention is given to patient preparation and selection. Emergency phone contact with the surgeon in case of complications is important to avoid unnecessary hospitalization.

Isolation of new ligands for orphan receptor MRGPRX1-hemorphins LVV-H7 and VV-H7.

The human MAS-related G protein-coupled receptor X1 (MRGPRX1) is a member of the GPCR family. The receptor is primate specific and expressed in the sensory neurons of dorsal root ganglion and trigeminal ganglion, where it is considered to be involved in the pain perception. The MRGPRX1 has unusual binding mechanism, as it is activated by several different ligands as well as several different fragments of precursor proteins. Thus, we hypothesize that it is activated by several unknown compounds as well since the receptor is still classified as orphan. Here, we describe the isolation of two novel endogenous ligands for the MRGPRX1 from human platelet preparation. The isolated ligands are hemoglobin ß-chain fragments, known members of the hemorphin family.

Mast cell degranulation via MRGPRX2 by isolated human albumin fragments.

BACKGROUND: Mast cells are important modulators of the human immune system via their release of several inflammatory mediators and proteases. The release can be activated by different pathways: the classical immunoglobulin E-dependent pathway and by the non-immunological immunoglobulin E-independent pathway. MAS-related G protein-coupled receptor X2 (MRGPRX2) is expressed in mast cells and it is one of the endogenous receptor responsible for the IgE-independent activation of human mast cell. The MRGPRX2 is classified as orphan receptor and unlike most GPCRs, the MRGPRX2 recognizes a wide range of basic molecules. Thus, there still might be several unknown ligands for the receptor. METHODS: MRGPRX2 activating peptides were isolated from human plasma using consecutive HPLC purification steps. The isolation process was monitored with MRGPRX2 transfected HEK 293 cells. The isolated peptides were sequenced by MS and synthetized. The synthetic peptides were used to determine degranulation of the human LAD 2 mast cell line by measuring ß-hexosaminidase release. RESULTS: Three endogenous MRGPRX2 activating peptides were isolated from human plasma. These peptides are identified as fragments of albumin. The isolated fragments activate MRGPRX2 and degranulate MRGPRX2 expressing LAD 2 cells in dose-dependent manner. CONCLUSIONS: The isolated basic peptides generated from human albumin are able to degranulate mast cells via the MRGPRX2. GENERAL SIGNIFICANCE: These endogenous albumin fragments, cleaved from albumin by mast cell secreted proteases, provide a possible pathway for self-perpetuating mast cell dependent inflammation.

Towards characterization of photo-excited electron transfer and catalysis in natural and artificial systems using XFELs.

The ultra-bright femtosecond X-ray pulses provided by X-ray Free Electron Lasers (XFELs) open capabilities for studying the structure and dynamics of a wide variety of biological and inorganic systems beyond what is possible at synchrotron sources. Although the structure and chemistry at the catalytic sites have been studied intensively in both biological and inorganic systems, a full understanding of the atomic-scale chemistry requires new approaches beyond the steady state X-ray crystallography and X-ray spectroscopy at cryogenic temperatures. Following the dynamic changes in the geometric and electronic structure at ambient conditions, while overcoming X-ray damage to the redox active catalytic center, is key for deriving reaction mechanisms. Such studies become possible by using the intense and ultra-short femtosecond X-ray pulses from an XFEL, where sample is probed before it is damaged. We have developed methodology for simultaneously collecting X-ray diffraction data and X-ray emission spectra, using an energy dispersive spectrometer, at ambient conditions, and used this approach to study the room temperature structure and intermediate states of the photosynthetic water oxidizing metallo-protein, photosystem II. Moreover, we have also used this setup to simultaneously collect the X-ray emission spectra from multiple metals to follow the ultrafast dynamics of light-induced charge transfer between multiple metal sites. A Mn-Ti containing system was studied at an XFEL to demonstrate the efficacy and potential of this method.

Leaf metallome preserved over 50 million years.

Large-scale Synchrotron Rapid Scanning X-ray Fluorescence (SRS-XRF) elemental mapping and X-ray absorption spectroscopy are applied here to fossil leaf material from the 50 Mya Green River Formation (USA) in order to improve our understanding of the chemistry of fossilized plant remains. SRS-XRF of fossilized animals has previously shown that bioaccumulated trace metals and sulfur compounds may be preserved in their original distributions and these elements can also act as biomarkers for specific biosynthetic pathways. Similar spatially resolved chemical data for fossilized plants is sparsely represented in the literature despite the multitude of other chemical studies performed. Here, synchrotron data from multiple specimens consistently show that fossil leaves possess chemical inventories consisting of organometallic and organosulfur compounds that: (1) map discretely within the fossils, (2) resolve fine scale biological structures, and (3) are distinct from embedding sedimentary matrices. Additionally, the chemical distributions in fossil leaves are directly comparable to those of extant leaves. This evidence strongly suggests that a significant fraction of the chemical inventory of the examined fossil leaf material is derived from the living organisms and that original bioaccumulated elements have been preserved in situ for 50 million years. Chemical information of this kind has so far been unknown for fossilized plants and could for the first time allow the metallome of extinct flora to be studied.

Effects of intra-arterial octreotide on pancreatic texture: a randomized controlled trial.

BACKGROUND AND AIMS: Octreotide is suggested to harden the pancreas, thus facilitating the construction of a pancreatic anastomosis and lowering the risk of postoperative fistula. We tested the hypothesis that intra-arterial application of octreotide in the gastroduodenal artery during pancreatectomy may increase pancreatic hardness. MATERIAL AND METHODS: A single-center, prospective, double-blinded, randomized controlled trial with parallel assignment was conducted. Patients planned for a pancreatoduodenectomy or a total pancreatectomy, who had a palpatory and durometer proven (<40 Shore units) soft pancreas, were assigned to receive intraoperatively either 5 mL 500µg octreotide or 5 mL 0.9% saline solution as a bolus injection in the gastroduodenal artery. Pancreatic hardness was measured before, early, and late after intervention. The investigator performing the durometer measurements and pathologist were masked to group assignment. The primary outcome was increased pancreatic hardness. Analysis was by intention to treat. This trial is registered at http://www.clinicaltrials.gov (ID NCT01400100). RESULTS: A total of 12 patients received octreotide and 13 received saline solution. Pancreatic hardness marginally increased in the octreotide group: 0.67 ± 2.3 Shore units, whereas it decreased in the control group: -2.15 ± 2.7 Shore units. The difference was statistically significant, p = 0.029 (95% confidence interval = -4.87 to -0.77). Histology did not find any correlate for this clinically irrelevant hardening effect. CONCLUSIONS: A single bolus application of octreotide did not deliver a clinically relevant increase in pancreatic hardness. Future studies on the hardening effect of octreotide should employ repeated or continuous preoperative administration of this drug.

A seven-crystal Johann-type hard x-ray spectrometer at the Stanford Synchrotron Radiation Lightsource.

We present a multicrystal Johann-type hard x-ray spectrometer (~5-18 keV) recently developed, installed, and operated at the Stanford Synchrotron Radiation Lightsource. The instrument is set at the wiggler beamline 6-2 equipped with two liquid nitrogen cooled monochromators--Si(111) and Si(311)--as well as collimating and focusing optics. The spectrometer consists of seven spherically bent crystal analyzers placed on intersecting vertical Rowland circles of 1 m of diameter. The spectrometer is scanned vertically capturing an extended backscattering Bragg angular range (88°-74°) while maintaining all crystals on the Rowland circle trace. The instrument operates in atmospheric pressure by means of a helium bag and when all the seven crystals are used (100 mm of projected diameter each), has a solid angle of about 0.45% of 4π sr. The typical resolving power is in the order of E/ΔE ~ 10,000. The spectrometer's high detection efficiency combined with the beamline 6-2 characteristics permits routine studies of x-ray emission, high energy resolution fluorescence detected x-ray absorption and resonant inelastic x-ray scattering of very diluted samples as well as implementation of demanding in situ environments.

Application of metal-coded affinity tags (MeCAT): absolute protein quantification with top-down and bottom-up workflows by metal-coded tagging.

As the quantification of peptides and proteins extends from comparative analyses to the determination of actual amounts, methodologies for absolute protein quantification are desirable. Metal-coded affinity tags (MeCAT) are chemical labels for peptides and proteins with a lanthanide-bearing chelator as a core. This modification of analytes with non-naturally occurring heteroelements adds the analytical possibilities of inductively coupled plasma mass spectrometry (ICPMS) to quantitative proteomics. We here present the absolute quantification of recombinantly expressed aprotinin out of its host cell protein background using two independent MeCAT methodologies. A bottom-up strategy employs labeling of primary amino groups on peptide level. Synthetic peptides with a MeCAT label which are externally quantified by flow injection analysis (FIA)-ICPMS serve as internal standard in nanoHPLC-ESI-MS/MS. In the top-down approach, protein is labeled on cysteine residues and separated by two-dimensional gel electrophoresis. Flow injection analysis of dissolved gel spots by ICPMS yields the individual protein amount via its lanthanide label content. The enzymatic determination of the fusion protein via its ß-galactosidase activity found 8.3 and 9.8 ng/µg (nanogram fusion protein per microgram sample) for batches 1 and 2, respectively. Using MeCAT values of 4.0 and 5.4 ng/µg are obtained for top-down analysis, while 14.5 and 15.9 ng/µg were found in the bottom-up analysis.

Can the new RCP R0/R1 classification predict the clinical outcome in ductal adenocarcinoma of the pancreatic head?

PURPOSE: According to the International Union Against Cancer (UICC), R1 is defined as the microscopic presence of tumor cells at the surface of the resection margin (RM). In contrast, the Royal College of Pathologists (RCP) suggested to declare R1 already when tumor cells are found within 1 mm of the RM. The aim of this study was to determine the significance of the RM concerning the prognosis of pancreatic ductal adenocarcinoma (PDAC). METHODS: From 2007 to 2009, 62 patients underwent a curative operation for PDAC of the pancreatic head. The relevance of R status on cumulative overall survival (OS) was assessed on univariate and multivariate analysis for both the classic R classification (UICC) and the suggestion of the RCP. RESULTS: Following the UICC criteria, a positive RM was detected in 8 %. Along with grading and lymph node ratio, R status revealed a significant impact on OS on univariate and multivariate analysis. Applying the suggestion of the RCP, R1 rate rose to 26 % resulting in no significant impact on OS in univariate analysis. CONCLUSIONS: Our study has shown that the RCP suggestion for R status has no impact on the prognosis of PDAC. In contrast, our data confirmed the UICC R classification of RM as well as N category, grading, and lymph node ratio as significant prognostic factors.

A high resolution and large solid angle x-ray Raman spectroscopy end-station at the Stanford Synchrotron Radiation Lightsource.

We present a new x-ray Raman spectroscopy end-station recently developed, installed, and operated at the Stanford Synchrotron Radiation Lightsource. The end-station is located at wiggler beamline 6-2 equipped with two monochromators-Si(111) and Si(311) as well as collimating and focusing optics. It consists of two multi-crystal Johann type spectrometers arranged on intersecting Rowland circles of 1 m diameter. The first one, positioned at the forward scattering angles (low-q), consists of 40 spherically bent and diced Si(110) crystals with 100 mm diameters providing about 1.9% of 4π sr solid angle of detection. When operated in the (440) order in combination with the Si (311) monochromator, an overall energy resolution of 270 meV is obtained at 6462.20 eV. The second spectrometer, consisting of 14 spherically bent Si(110) crystal analyzers (not diced), is positioned at the backward scattering angles (high-q) enabling the study of non-dipole transitions. The solid angle of this spectrometer is about 0.9% of 4π sr, with a combined energy resolution of 600 meV using the Si (311) monochromator. These features exceed the specifications of currently existing relevant instrumentation, opening new opportunities for the routine application of this photon-in/photon-out hard x-ray technique to emerging research in multidisciplinary scientific fields, such as energy-related sciences, material sciences, physical chemistry, etc.

[Indications for laparoscopic pancreas operations: results of a consensus conference and the previous laparoscopic pancreas register]. / Indikation zur laparoskopischen Pankreasoperation: Ergebnisse einer Konsensuskonferenz und des bisherigen laparoskopischen Pankreasregisters.

Laparoscopic pancreatic surgery is not common practice in Germany and is only carried out in approximately 20 clinics but with an increasing trend. The reasons for this are manifold, such as the current selection of patients and both skills in laparoscopic and pancreatic surgery are necessary to perform this operation safely. In 2008 a registry called "Laparoscopic pancreatic surgery" was implemented to collect enough data in Germany to find out whether the resection is safe, feasible and beneficial for the patient.For further development of new laparoscopic techniques new data is needed. A group of experts performing laparoscopic pancreatic surgery in Germany supplied their data for the German registry for laparoscopic pancreatic resection and a consensus conference about the indications became necessary. This consensus conference discussed in particular the indications for laparoscopic pancreatic resection. A consensus was found by all members of the conference utilizing currently available evidence-based data.It was suggested that all data of laparoscopic pancreatic surgery should be evaluated in the German Registry. A consensus was made which diseases were either suitable for laparoscopic resection or not suitable or suitable in selected cases.

Wide-angle X-ray diffraction and molecular dynamics study of medium-range order in ambient and hot water.

We have developed wide-angle X-ray diffraction measurements with high energy-resolution and accuracy to study water structure at three different temperatures (7, 25 and 66 °C) under normal pressure. Using a spherically curved Ge crystal an energy resolution better than 15 eV has been achieved which eliminates influence from Compton scattering. The high quality of the data allows for a reliable Fourier transform of the experimental data resolving shell structure out to ~12 Å, i.e. 5 hydration shells. Large-scale molecular dynamics (MD) simulations using the TIP4P/2005 force-field reproduce excellently the experimental shell-structure in the range 4-12 Å although less agreement is seen for the first peak in the intermolecular pair-correlation function (PCF). The Shiratani-Sasai Local Structure Index [J. Chem. Phys. 104, 7671 (1996)] identifies a tetrahedral minority giving the intermediate-range oscillations in the O-O PCF and a disordered majority providing a more featureless background in this range. The current study supports the proposal that the structure of liquid water, even at high temperatures, can be described in terms of a two-state fluctuation model involving local structures related to the high-density and low-density forms of liquid water postulated in the liquid-liquid phase transition hypothesis.

Trace metals as biomarkers for eumelanin pigment in the fossil record.

Well-preserved fossils of pivotal early bird and nonavian theropod species have provided unequivocal evidence for feathers and/or downlike integuments. Recent studies have reconstructed color on the basis of melanosome structure; however, the chemistry of these proposed melanosomes has remained unknown. We applied synchrotron x-ray techniques to several fossil and extant organisms, including Confuciusornis sanctus, in order to map and characterize possible chemical residues of melanin pigments. Results show that trace metals, such as copper, are present in fossils as organometallic compounds most likely derived from original eumelanin. The distribution of these compounds provides a long-lived biomarker of melanin presence and density within a range of fossilized organisms. Metal zoning patterns may be preserved long after melanosome structures have been destroyed.

Infrared mapping resolves soft tissue preservation in 50 million year-old reptile skin.

Non-destructive Fourier Transform InfraRed (FTIR) mapping of Eocene aged fossil reptile skin shows that biological control on the distribution of endogenous organic components within fossilized soft tissue can be resolved. Mapped organic functional units within this approximately 50 Myr old specimen from the Green River Formation (USA) include amide and sulphur compounds. These compounds are most probably derived from the original beta keratin present in the skin because fossil leaf- and other non-skin-derived organic matter from the same geological formation do not show intense amide or thiol absorption bands. Maps and spectra from the fossil are directly comparable to extant reptile skin. Furthermore, infrared results are corroborated by several additional quantitative methods including Synchrotron Rapid Scanning X-Ray Fluorescence (SRS-XRF) and Pyrolysis-Gas Chromatography/Mass Spectrometry (Py-GC/MS). All results combine to clearly show that the organic compound inventory of the fossil skin is different from the embedding sedimentary matrix and fossil plant material. A new taphonomic model involving ternary complexation between keratin-derived organic molecules, divalent trace metals and silicate surfaces is presented to explain the survival of the observed compounds. X-ray diffraction shows that suitable minerals for complex formation are present. Previously, this study would only have been possible with major destructive sampling. Non-destructive FTIR imaging methods are thus shown to be a valuable tool for understanding the taphonomy of high-fidelity preservation, and furthermore, may provide insight into the biochemistry of extinct organisms.

Complicated sigmoid diverticulitis--Hartmann's procedure or primary anastomosis?

INTRODUCTION: Diverticular disease of the colon is a common condition in developed countries. For perforated diverticulitis Hartmann's procedure is a safe and quick treatment option. But intestinal restoration needs further interventions. This leads to high complication rates and cost. Therefore a critical evaluation of surgical treatment options is necessary. METHODS: During a period of 18 months 88 patients underwent surgical resection for diverticulitis. Forty patients had emergency surgery. Among those a primary anastomosis was performed in 21 patients. The other 19 patients had interval colostomy. Among 21 patients with primary anastomosis major complications occurred in two patients, vs. twelve in patients with Hartmann's operation (p = 0.03). In the Hartmann group eight patients had major general complications, vs. one patient in the group with primary anastomosis (p = 0.06). The mean hospital stay was 38 days after Hartmann's procedure, vs. 13 days for patients with primary anastomosis (p < 0.01). CONCLUSION: In emergency surgery for complicated diverticulitis primary anastomosis is not associated with an increased postoperative morbidity. A primary anastomosis reduces the need for further surgical interventions and complex re-operations. Thus, an overall reduction of morbidity, cost, complication rate and hospital stay is possible. Therefore this technique is advantageous for patients and hospitals.

Archaeopteryx feathers and bone chemistry fully revealed via synchrotron imaging.

Evolution of flight in maniraptoran dinosaurs is marked by the acquisition of distinct avian characters, such as feathers, as seen in Archaeopteryx from the Solnhofen limestone. These rare fossils were pivotal in confirming the dinosauria-avian lineage. One of the key derived avian characters is the possession of feathers, details of which were remarkably preserved in the Lagerstätte environment. These structures were previously simply assumed to be impressions; however, a detailed chemical analysis has, until now, never been completed on any Archaeopteryx specimen. Here we present chemical imaging via synchrotron rapid scanning X-ray fluorescence (SRS-XRF) of the Thermopolis Archaeopteryx, which shows that portions of the feathers are not impressions but are in fact remnant body fossil structures, maintaining elemental compositions that are completely different from the embedding geological matrix. Our results indicate phosphorous and sulfur retention in soft tissue as well as trace metal (Zn and Cu) retention in bone. Other previously unknown chemical details of Archaeopteryx are also revealed in this study including: bone chemistry, taphonomy (fossilization process), and curation artifacts. SRS-XRF represents a major advancement in the study of the life chemistry and fossilization processes of Archaeopteryx and other extinct organisms because it is now practical to image the chemistry of large specimens rapidly at concentration levels of parts per million. This technique has wider application to the archaeological, forensic, and biological sciences, enabling the mapping of "unseen" compounds critical to understanding biological structures, modes of preservation, and environmental context.

The inhomogeneous structure of water at ambient conditions.

Small-angle X-ray scattering (SAXS) is used to demonstrate the presence of density fluctuations in ambient water on a physical length-scale of approximately 1 nm; this is retained with decreasing temperature while the magnitude is enhanced. In contrast, the magnitude of fluctuations in a normal liquid, such as CCl(4), exhibits no enhancement with decreasing temperature, as is also the case for water from molecular dynamics simulations under ambient conditions. Based on X-ray emission spectroscopy and X-ray Raman scattering data we propose that the density difference contrast in SAXS is due to fluctuations between tetrahedral-like and hydrogen-bond distorted structures related to, respectively, low and high density water. We combine our experimental observations to propose a model of water as a temperature-dependent, fluctuating equilibrium between the two types of local structures driven by incommensurate requirements for minimizing enthalpy (strong near-tetrahedral hydrogen-bonds) and maximizing entropy (nondirectional H-bonds and disorder). The present results provide experimental evidence that the extreme differences anticipated in the hydrogen-bonding environment in the deeply supercooled regime surprisingly remain in bulk water even at conditions ranging from ambient up to close to the boiling point.

Spectroscopic characterization of microscopic hydrogen-bonding disparities in supercritical water.

The local hydrogen-bonding environment in supercritical water (380 degrees C, 300 bars, density 0.54 gcm3) was studied by x-ray Raman scattering at the oxygen K edge. The spectra are compared to those of the gas phase, liquid surface, bulk liquid, and bulk ice, as well as to calculated spectra. The experimental model systems are used to assign spectral features and to quantify specific local hydrogen-bonding situations in supercritical water. The first coordination shell of the molecules is characterized in more detail with the aid of the calculations. Our analysis suggests that approximately 65% of the molecules in supercritical water are hydrogen bonded in configurations that are distinctly different from those in liquid water and ice. In contrast to liquid water the bonded molecules in supercritical water have four intact hydrogen bonds and in contrast to ice large variations of bond angles and distances are observed. The remaining approximately 35% of the molecules exhibit two free O-H bonds and are thus either not involved in hydrogen bonding at all or have one or two hydrogen bonds on the oxygen side. We determine an average O-O distance of 3.1+/-0.1 A in supercritical water for the H bonded molecules at the conditions studied here. This and the corresponding hydrogen bond lengths are shown to agree with neutron- and x-ray-diffraction data at similar conditions. Our results on the local hydrogen-bonding environment with mainly two disparate hydrogen-bonding configurations are consistent with an extended structural model of supercritical water as a heterogeneous system with small patches of bonded molecules in various tetrahedral configurations and surrounding nonbonded gas-phase-like molecules.