Cation-specific interactions of protein surface charges in dilute aqueous salt solutions: a combined study using dielectric relaxation spectroscopy and Raman spectroscopy.

We exploited glycine as a zwitterionic model system to experimentally probe the cation specific interaction of protein surface charges in dilute (≤0.25 mol L-1) aqueous solutions of four biologically relevant inorganic salts, NaCl, KCl, MgCl2 and CaCl2, via dielectric relaxation spectroscopy (DRS) and Raman spectroscopy. Glycine is the simplest building block of proteins and it exposes the same charged groups (carboxylate and ammonium) to the solvent that dominate the protein-water interface. As a counter ion, we selected Cl- due to its biological importance. For all systems, we performed simultaneous fitting of the real (ε') and imaginary (ε″) parts of the dielectric functions, assuming a multimodal relaxation model, obtained from concentration dependent dielectric measurements at ∼293 K. We observe a reduction of the dielectric amplitude for the glycine relaxation while the corresponding time constant shows only small (<7%) deviations compared to aqueous glycine solutions. We propose that the observed reduction in dielectric amplitude is due to a reduction of the effective dipole moment (µeff) of zwitterionic glycine caused by the interaction of glycine with the ion even at very low (0.05 M) salt concentrations. The interaction between divalent metal ions and zwitterionic glycine is increased compared to the monovalent cation-zwitterion interaction; a finding that is also supported by Raman spectroscopy. Our combined dielectric relaxation and Raman spectroscopic study indicates that ion-glycine interactions are weak and mediated by the solvent. Cation-specificity of protein surface charges is also observed in dilute salt solutions (≤0.25 mol L-1), where electrostatic interactions dominate.

Pilot study demonstrates that placental histology can provide an additional tool for diagnosing early-onset neonatal sepsis.

AIM: We explored whether placental histology could help to diagnose early-onset neonatal sepsis (EONS), guide clinical decision-making 48 hours after birth and reduce antibiotic use. METHODS: This study comprised 109 infants born at less than 32 weeks of gestation, who were admitted to the neonatal intensive care unit of Isala, Zwolle, The Netherlands, between January 2013 and December 2013. EONS was defined as clinical symptoms plus raised serial C-reactive protein (CRP) >10 mg/L and a positive (proven EONS) or a negative (suspected EONS) blood culture. Placentas were studied for a histological inflammatory response and scored according to Redline's criteria. RESULTS: A histological inflammatory response was seen in 15/88 (17%) placentas and this occurred significantly more often in infants with a high suspicion of EONS (p < 0.05). No histological inflammatory response was seen if maternal risk factors for EONS were absent, despite a raised CRP level. Based on placental histology, the duration of antibiotic therapy was reduced from more than five days to 48 hours in 20/27 infants (74%). CONCLUSION: Histological examination of the placenta helped to diagnose EONS and guide clinical decision-making 48 hours after birth and led to a clinically relevant reduction in antibiotic use.

Consequence of restaging after neoadjuvant treatment for locally advanced rectal cancer.

BACKGROUND: Locally advanced rectal cancer is customarily treated with neoadjuvant chemoradiotherapy (CRT) followed by a total mesorectal excision. During the course of CRT, previously non-detectable distant metastases can appear. Therefore, a restaging CT scan of the chest and abdomen was performed prior to surgery. The aim of this study was to determine the frequency of a change in treatment strategy after this restaging CT scan. METHODS: Patients treated with neoadjuvant CRT for locally advanced rectal cancer between January 2003 and July 2013 were included retrospectively. To determine the value of the restaging CT scan, the surgical treatment as planned before CRT was compared with the treatment ultimately received. RESULTS: A total of 153 patients (91 male) were eligible, and median age was 62 (32-82) years. The restaging CT scan revealed the presence of distant metastases in 19 patients (12.4, 95 % confidence interval [CI] 7.0-17.8). In 17 patients (11.1, 95 % CI 6.1-16.1), a change in treatment strategy occurred due to the detection of metastases with a restaging CT scan. CONCLUSION: A restaging CT scan after completion of neoadjuvant CRT may detect newly developed metastases and consequently alter the initial treatment strategy. This study demonstrated the added value of the restaging CT scan prior to surgery.

Graphene multilayer as nanosized optical strain gauge for polymer surface relief gratings.

In this paper, we show how graphene can be utilized as a nanoscopic probe in order to characterize local opto-mechanical forces generated within photosensitive azobenzene containing polymer films. Upon irradiation with light interference patterns, photosensitive films deform according to the spatial intensity variation, leading to the formation of periodic topographies such as surface relief gratings (SRG). The mechanical driving forces inscribing a pattern into the films are supposedly fairly large, because the deformation takes place without photofluidization; the polymer is in a glassy state throughout. However, until now there has been no attempt to characterize these forces by any means. The challenge here is that the forces vary locally on a nanometer scale. Here, we propose to use Raman analysis of the stretching of the graphene layer adsorbed on top of polymer film under deformation in order to probe the strength of the material transport spatially resolved. With the well-known mechanical properties of graphene, we can obtain lower bounds on the forces acting within the film. Upon the basis of our experimental results, we can deduce that the internal pressure in the film due to grating formation can exceed 1 GPa. The graphene-based nanoscopic gauge opens new possibilities to characterize opto-mechanical forces generated within photosensitive polymer films.

Local chemical composition of nanophase-separated polymer brushes.

Using scattering scanning nearfield infrared microscopy (s-SNIM), we have imaged the nanoscale phase separation of mixed polystyrene-poly(methyl methacrylate) (PS-PMMA) brushes and investigated changes in the top layer as a function of solvent exposure. We deduce that the top-layer of the mixed brushes is composed primarily of PMMA after exposure to acetone, while after exposure to toluene this changes to PS. Access to simultaneously measured topographic and chemical information allows direct correlation of the chemical morphology of the sample with topographic information. Our results demonstrate the potential of s-SNIM for chemical mapping based on distinct infrared absorption properties of polymers with a high spatial resolution of 80 nm × 80 nm.

Wrapping Up Hydrophobic Hydration: Locality Matters.

Water, being the universal solvent, acts as a competing agent in fundamental processes, such as folding, aggregation or biomolecular recognition. A molecular understanding of hydrophobic hydration is of central importance to understanding the subtle free energy differences, which dictate function. Ab initio and classical molecular dynamics simulations yield two distinct hydration water populations in the hydration shell of solvated tert-butanol noted as "HB-wrap" and "HB-hydration2bulk". The experimentally observed hydration water spectrum can be dissected into two modes, centered at 164 and 195 cm-1. By comparison to the simulations, these two bands are attributed to the "HB-wrap" and "HB-hydration2bulk" populations, respectively. We derive a quantitative correlation between the population in each of these two local water coordination motifs and the temperature dependence of the solvation entropy. The crossover from entropy to enthalpy dominated solvation at elevated temperatures, as predicted by theory and observed experimentally, can be rationalized in terms of the distinct temperature stability and thermodynamic signatures of "HB-wrap" and "HB-hydration2bulk".

Integration of a rapid scanning technique into THz time-domain spectrometers for nonlinear THz spectroscopy measurements.

We have implemented a rapid scanning technique into THz time-domain spectrometers using an oscillating frictionless delay line, especially adapted for nonlinear THz experiments. Thereby we were able to increase the dynamic range of THz measurements in the frequency range from 40 to 200 cm-1 by up to 24 dB and reduce the scanning time by up to a factor of 200. We report here test measurements on TDS-setups at repetition rates of 80 MHz and 5 kHz. The dynamic range exceeds 64 dB, which allows to record even small changes in the THz absorption upon optical excitation by a THz probe, covering the frequency range of the intermolecular modes and the phonon bands. We demonstrate the potential of this technique for optical-pump THz-probe experiments using a 70 µm thick high-resistivity silicon, excited by 400 nm, ∼50 fs pulses as a sample.

Aggression in BALB/cJ mice is differentially predicted by the volumes of anterior and midcingulate cortex.

Anterior cingulate cortex (ACC) and midcingulate cortex (MCC) have been implicated in the regulation of aggressive behaviour. For instance, patients with conduct disorder (CD) show increased levels of aggression accompanied by changes in ACC and MCC volume. However, accounts of ACC/MCC changes in CD patients have been conflicting, likely due to the heterogeneity of the studied populations. Here, we address these discrepancies by studying volumetric changes of ACC/MCC in the BALB/cJ mouse, a model of aggression, compared to an age- and gender-matched control group of BALB/cByJ mice. We quantified aggression in BALB/cJ and BALB/cByJ mice using the resident-intruder test, and related this to volumetric measures of ACC/MCC based on Nissl-stained coronal brain slices of the same animals. We demonstrate that BALB/cJ behave consistently more aggressively (shorter attack latencies, more frequent attacks, anti-social biting) than the control group, while at the same time showing an increased volume of ACC and a decreased volume of MCC. Differences in ACC and MCC volume jointly predicted a high amount of variance in aggressive behaviour, while regression with only one predictor had a poor fit. This suggests that, beyond their individual contributions, the relationship between ACC and MCC plays an important role in regulating aggressive behaviour. Finally, we show the importance of switching from the classical rodent anatomical definition of ACC as cingulate area 2 and 1 to a definition that includes the MCC and is directly homologous to higher mammalian species: clear behaviour-related differences in ACC/MCC anatomy were only observed using the homologous definition.

Long-range influence of carbohydrates on the solvation dynamics of water--answers from terahertz absorption measurements and molecular modeling simulations.

We present new terahertz (THz) spectroscopic measurements of solvated sugars and compare the effect of two disaccharides (trehalose and lactose) and one monosaccharide (glucose) with respect to the solute-induced changes in the sub-picosecond network dynamics of the hydration water. We found that the solute affects the fast collective network motions of the solvent, even beyond the first solvation layer. For all three carbohydrates, we find an increase of 2-4% in the THz absorption coefficient of the hydration water in comparison to bulk water. Concentration-dependent changes in the THz absorption between 2.1 and 2.8 THz of the solute-water mixture were measured with a precision better than 1% and were used to deduce a dynamical hydration shell, which extends from the surface up to 5.7 +/- 0.4 and 6.5 +/- 0.9 A for the disaccharides lactose and trehalose, respectively, and 3.7 +/- 0.9 A for the glucose. This exceeds the values for the static hydration shell as determined, for example, by scattering, where the long-range structure was found to be not significantly affected by the solute beyond the first hydration shell. When comparing all three carbohydrates, we found that the solute-induced change in the THz absorption depends on the product of molar concentration of the solute and the number of hydrogen bonds between the carbohydrate and water molecules. We can conclude that the long-range influence on the sub-picosecond collective water network motions of the hydration water is directly correlated with the average number of hydrogen bonds between the molecule and adjacent water molecules for carbohydrates. This implies that monosaccharides have a smaller influence on the surrounding water molecules than disaccharides. This could explain the bioprotection mechanism of sugar-water mixtures, which has been found to be more effective for disaccharides than for monosaccharides.

The opto-locomotor reflex as a tool to measure sensitivity to moving random dot patterns in mice.

We designed a method to quantify mice visual function by measuring reflexive opto-locomotor responses. Mice were placed on a Styrofoam ball at the center of a large dome on the inside of which we projected moving random dot patterns. Because we fixed the heads of the mice in space and the ball was floating on pressurized air, locomotion of the mice was translated to rotation of the ball, which we registered. Sudden onsets of rightward or leftward moving patterns caused the mice to reflexively change their running direction. We quantified the opto-locomotor responses to different pattern speeds, luminance contrasts, and dot sizes. We show that the method is fast and reliable and the magnitude of the reflex is stable within sessions. We conclude that this opto-locomotor reflex method is suitable to quantify visual function in mice.

Comparison of pregnancy course and outcome with color and radiographic angiography of the placenta in a monochorionic triplet pregnancy.

A patient with spontaneous monochorionic (MC) triamniotic triplet pregnancy developed symptoms of feto-fetal-transfusion-syndrome (FFTS) at 18 weeks of gestation with one donor (oligohydramnios) and one receptor (polyhydramnios) triplet. The patient "received" a diet enriched with proteins. Amniotic-fluid volume returned to normal after 24 weeks. At 32 weeks, a Cesarean section was performed due to intra-uterine growth restriction of the donor triplet. Post partum color injection and computer angiograms showed arterio-arterial (AA) anastomoses between all triplets. Deep arterio-venous (AV) anastomoses between the two triplets who had demonstrated with oligo- and polyhydramnios between 18 and 26 weeks were revealed by computer angiography that were not seen by placental color angiogram. Detailed analysis of placental vascular communications by use of color injection angiogram of the chorionic plate and computer angiogram demonstrating deep anastomoses beneath the chorionic plate helps to understand the individual pathophysiology and clinical course in patients with FFTS, which is even more complex in MC triplet compared to MC twin pregnancies.

A novel set-up to investigate the low-frequency spectra of aqueous solutions at high hydrostatic pressure.

We present a novel setup to investigate the low frequency (THz/FIR) spectra of an aqueous solution under high hydrostatic pressure (HHP). By integration of a diamond anvil cell into a THz Fourier transform spectrometer, we are able to record the absorption of bulk water in the pressure range from 1 bar to 10 kbar. The difference in intensity can directly be compared to the difference in extinction coefficients. The spectroscopic data reveal a blue shift of the H-bond stretch vibration at 180 cm-1, which is evidence of changes in the H-bond network dynamics.

Long-term follow-up (12 to 35 weeks) after dynamic cardiomyoplasty.

OBJECTIVES: To obtain information on the long-term effects of dynamic cardiomyoplasty on hemodynamics and muscle histology, this surgical method was evaluated in goats. BACKGROUND: Dynamic cardiomyoplasty has been introduced as a new method to treat patients with severe cardiac failure. METHODS: In 24 goats, the left latissimus dorsi muscle was wrapped around the heart. The muscle was then subjected to progressive electrical stimulation. In 16 goats, invasive transesophageal Doppler echocardiographic measurements and histologic evaluation of the latissimus dorsi muscle were performed at > or = 12 weeks after the wrapping. RESULTS: Only two goats showed an increase in aortic and left and right ventricular pressures concomitant with increased aortic flow during latissimus dorsi muscle stimulation both before and after induction of cardiac failure using imipramine. This was accompanied by a preserved latissimus dorsi muscle structure and nearly complete transformation to type I muscle fibers. The remaining 14 goats showed extensive lipomatosis in the latissimus dorsi muscle, with severe intimal hyperplasia and proliferation of smooth muscle cells in the walls of the thoracodorsal artery and its branches. An increase in endoneural and endomysial connective tissue was observed, with some goats showing destroyed nerve branches near the electrodes. These findings differed from those observed after long-term electrical stimulation of goat latissimus dorsi muscle in situ. CONCLUSIONS: Dynamic cardiomyoplasty is of use in the treatment of severe heart failure if the histologic structure of the wrapped latissimus dorsi muscle remains intact. Long-term results in goats suggest that the current approach used in dynamic cardiomyoplasty may lead to deterioration of the wrapped muscle.

Immunohistochemical detection of very recent myocardial infarctions in humans with antibodies against heart-type fatty acid-binding protein.

Depletion of human heart-type fatty acid-binding protein (H-FABP) from cardiomyocytes in infarcted areas with varying postinfarction intervals was studied in 25 autopsy cases. In 23 autopsy cases myocardial infarction was the clinical diagnosis; 2 cases were noncardiac deaths and served as controls. Formaldehyde-fixed and paraffin-embedded myocardial tissue sections were stained immunohistochemically with antibodies to human H-FABP. H-FABP immunohistochemical findings were compared with those from conventional nitroblue tetrazolium (NBT) macroenzyme staining and conventional histochemical hematoxylin-eosin staining of sections of the same infarctions. In all cases of infarction confirmed by NBT staining, decreased or absent H-FABP immunostaining was observed. In 12 cases of clinically diagnosed infarction, H-FABP depletion could be demonstrated in areas that showed normal NBT staining. These findings strongly suggest that immunohistochemical staining with antibodies to H-FABP can confirm the clinical diagnosis or suspicion of early myocardial infarction. In both control cases no depletion of H-FABP was observed in cardiomyocytes in different myocardial tissue sections, and macroenzyme staining with NBT was normal. We conclude that H-FABP immunostaining may detect very recent ischemia/ infarction in human myocardium and can be applied in routine autopsy pathology.

Changes in canine latissimus dorsi muscle during 24 wk of continuous electrical stimulation.

To study functional, structural, and biochemical adaptations to electrical stimulation of striated muscle in a large animal, the canine latissimus dorsi (LD) muscle was conditioned continuously for 24 wk with an increasing number of pulse bursts (burst duration 250 ms, burst frequency 30 Hz). Force measurements in vivo after 12 wk showed a significant decrease in the ripple, the ratio of interstimulus to peak force amplitude, from 0.94 +/- 0.03 to 0.13 +/- 0.08 (SE; n = 8, P less than 0.05), indicating reduction in contractile speed. Also the steep part of the force-frequency relation shifted to lower frequencies. A significant change in fiber-type composition was seen with both enzyme- and immunohistochemistry, manifested by an increase of type I fibers from 29.5 +/- 2.9 to 83 +/- 8% (SE; n = 8, P less than 0.05). During this period a transient rise in the number of type IIc/Ic fibers (from 3 to 10%) was seen. In the stimulated muscle, capillary-to-fiber ratio increased from 1.9 +/- 0.4 to 2.7 +/- 0.1 (P less than 0.05). A significant increase in mitochondrial volume was also seen, especially in the peripheral part of the fiber. Both creatine kinase and lactate dehydrogenase revealed a significant decline in activity within 12 wk. At the same time a shift in lactate dehydrogenase-isozyme pattern was observed toward the cardiac composition. No additional changes occurred after 12 wk of stimulation, indicating that conversion of the canine LD muscle was complete within this period.

Histopathologic validation of intracoronary ultrasound imaging.

The purpose of this study was to validate intracoronary ultrasound imaging by correlation with histologic examination. In this in-vitro study of pressure-perfused human coronary arteries, 104 matching intracoronary ultrasound imaging images and histologic cross-sections from 12 hearts were compared to determine the diagnostic accuracy of 30 MHz commercially available intracoronary ultrasound imaging. For lipid deposits, sensitivity was 46% and specificity 97%. The smallest lipid deposit that was visualized measured 0.25 mm in axial diameter on histologic study. For calcific deposits, sensitivity was 77% and specificity 100%. The smallest calcific deposit that was visualized measured 0.25 mm in axial diameter on histologic examination. Atherosclerotic intimal thickening could not be distinguished qualitatively or quantitatively from nonatherosclerotic intimal thickening unless there were localized deposits of lipids or calcium. Intracoronary ultrasound imaging is accurate in detecting lipid and calcium deposits. Intimal thickening in intracoronary ultrasound imaging images does not prove the presence of atherosclerosis.

Evaluation of gracilis muscle transposition for fecal incontinence with magnetic resonance imaging.

Ten consecutive patients with incapacitating fecal incontinence were treated with 'anal dynamic graciloplasty' (transposition of the gracilis muscle around the anal canal and implantation of intramuscular electrodes connected with an implanted pulse generator, 6 weeks later) to achieve continence. We measured the gracilis muscle diameter immediately after transposition and before implantation of the stimulation device. It was found that gracilis diameter decreased from 12 (5 days after transposition) to 8 mm, 6 weeks later (mean decrease: 4 mm (95% confidence interval 3.6), n = 10, P < 0.05). In addition, morphology demonstrated a decrease of both Type I and Type II muscle fiber diameter and an increase in endomysial collagen. Despite this decrease in muscle (and muscle fiber) diameter, electrical stimulation of the transposed gracilis muscle increased the pressure into the anal canal from 37 to 55 mmHg (mean increase: 17 mmHg (95% confidence interval 6.29), P < 0.05). Fecal continence was achieved in seven (70%) of these patients. Further analysis revealed no correlations between reduction of the gracilis muscle diameter before implantation of the stimulation device and clinical outcome in terms of achieved continence and/or anal canal pressures. MRI is an excellent method to demonstrate the shape of gracilis muscle after transposition. However, the size of transposed gracilis muscle is not associated with the functional outcome.

Characterization of Cobblestone mitral valve interstitial cells.

Mitral valve interstitial cells (MVICs) are important in the structure and function of the valve. In a cell culture system that used explants to harvest MVICs, we found that after several passages, some of the cultures at confluence formed a monolayer of cobblestone-shaped cells (CB type). The rest of the cultures were, however, different, consisting of elongated cells (EL type) that formed overlapping orthogonal patterns. When compared with the MVICs-EL, cells from cultures of MVICs-CB were less adherent to a plastic substratum initially than were MVICs-EL and were more sensitive to trypsinization following culture for 3 to 7 days. Ultrastructurally, the cultures of MVICs-CB showed prominent Golgi's complexes and endoplasmic reticulum and fewer microfilaments when compared with those of MVICs-EL. Most strikingly, the CB cells contained much less alpha-smooth-muscle cell actin compared with that of EL cells. The MVICs-CB were not endothelial cells as they did not show the characteristic dense peripheral band that was present in endothelial cells in a confluent monolayer and showed only trace amounts of fluorescence when incubated with 1,1-dioctadecyl 1-3,3,3,3 tetramethyl-indocarbocyanine-percholate (Dil-)--acetylated low-density lipoprotein compared with large amounts of fluorescence with endothelial cells. Cells that were morphologically similar to MVICs-CB were present in the in vivo valve as well and also formed small islands of cells even in the primary cultures that grew out of the explant. The findings supported the hypothesis that MVICs-CB represent a distinct phenotype of the MVICs that is different from that of the MVICs-EL and is not that of either endothelial or medial smooth-muscle cells.

De la Chapelle dysplasia (atelosteogenesis type II): case report and review of the literature [corrected].

We report a male infant with de la Chapelle dysplasia (atelosteogenesis type II), a skeletal dysplasia characterized by severe shortening of the long bones, deficient ossification of distinct parts of the skeleton, cleft palate and neonatal death from asphyxia. This is a rare condition with only 10 patients described in the literature. We report the clinical, radiographical and histopathological data and summarize the data on the total of 11 patients. Differential diagnosis with diastrophic dysplasia and atelosteogenesis (type I) is discussed. On clinical and histological grounds we hypothesize that de la Chapelle dysplasia and diastrophic dysplasia are closely related. The mode of inheritance is autosomal recessive.

[Anal dynamic plastic surgery of the gracilis muscle; a new surgical technique for the treatment of fecal incontinence]. / Anale dynamische M. gracilis-plastiek; een nieuwe chirurgische techniek voor de behandeling van fecale incontinentie.

OBJECTIVE: To study the clinical outcome of anal dynamic graciloplasty (gracilis muscle transposition and implantation of electric stimulation device) in a consecutive series of 12 patients. DESIGN: Prospective study. SETTING: Maastricht University Hospital. PATIENTS AND METHODS: Twelve patients with incapacitating faecal incontinence were treated using anal dynamic graciloplasty because they were not amenable to other medical management. The data were analysed with emphasis on the clinical outcome, anal manometry, and gracilis muscle composition before and after electric stimulation. Results were considered significant if p < or = 0.05, using the paired Student's t-test. RESULTS: Eight patients achieved complete faecal continence, one patient still has a previously constructed colostomy and in three patients no faecal incontinence could be achieved, due to infections. Median follow-up is now over 18 months (range 16 weeks-5 years). Anal manometry demonstrated an anal pressure increased from 39 mmHg (without stimulation) to 66 mmHg with electric stimulation (mean increase 27 mmHg (CI: 19-35; n = 12, p < 0.01). Gracilis muscle composition showed an increase of type I relatively fatigue-resistant fibres, capable of prolonged contractions, from 45% before stimulation to 64% afterwards (mean increase 19% (CI: 14-21; n = 8, p < 0.01). CONCLUSION: Dynamic graciloplasty is capable of replacing the function of damaged or absent anal sphincters. The construction of a colostomy in patients with incapacitating faecal incontinence can be avoided.