On the use of O2 spin-rotation lines for elevation angle calibration of atmospheric thermal emission spectra

The magnetic dipole-allowed spin-rotation lines of O₂ potentially can be used for pointing calibration and confirmation of float altitude for far infrared spectra of the stratosphere obtained from balloon platforms. We demonstrate that current deficiencies in the spectroscopic database, particularly the air pressure-broadening coefficients, severely limit the capability of deriving useful pointing calibrations, and that precise pressure-broadening measurements for the transitions actually used in calibration are needed to improve this capability.     

Antibiotic susceptibility testing (agar disk diffusion and agar dilution) of clinical isolates of Enterococcus faecalis and E. faecium: comparison of Mueller-Hinton, Iso-Sensitest, and Wilkins-Chalgren agar media

Forty-two isolates of Enterococcus faecalis and 56 isolates of Enterococcus faecium, including 8 vancomycin-resistant strains, were examined for comparative susceptibility to 27 antimicrobial drugs with the agar dilution method, employing Mueller-Hinton (MHA), Iso-Sensitest (ISTA), and Wilkins-Chalgren (WCA) agar. The Bauer-Kirby agar disk diffusion method was used to comparatively test 24 of the agents in parallel. The enterococci yielded better growth on ISTA and WCA. However, WCA completely antagonized co-trimoxazole and, though less, fosfomycin. Importantly, WCA slightly reduced the activities of teicoplanin (minimal inhibitory concentrations, MICs, raised up to twofold) and vancomycin (MICs raised two- to fourfold) against enterococci and staphylococcal quality control strains. Therefore, WCA was judged unsuitable for susceptibility testing of enterococci. For E. faecalis no discrepancies between agar dilution MICs and inhibition zone diameters were encountered with augmentin, ampicillin, ampicillin-sulbactam, chloramphenicol, mupirocin, oxacillin, teicoplanin, and co-trimoxazole. Overall, MHA yielded fewer very major (category I) and major (category II) discrepancies than ISTA. However, numerous minor (category III), slight (category IV), minimal (category V), and/or negligible (category VI) discrepancies were encountered with ciprofloxacin, doxycycline, erythromycin, fosfomycin, fusidic acid, meropenem, ofloxacin and rifampin. With respect to E. faecium, only cefotaxime, mupirocin, oxacillin, and teicoplanin yielded nondiscrepant results. Several very major (I) and major (II) discrepancies were observed with augmentin, ampicillin, ampicillin-sulbactam, doxycycline, fusidic acid, imipenem, and penicillin G. Minor discrepancies (categories III-VI) were particularly numerous with augmentin, chloramphenicol, ciprofloxacin, doxycycline, and piperacillin. The largest numbers of negligible (VI) discrepancies were noted with fosfomycin, fusidic acid, and ofloxacin. It is recommended to test one cephalosporin (cefuroxime or the like) in parallel for educational purposes and to exclude fosfomycin, fusidic acid, and rifampin from test batteries because of the wide scatter of test results. The large number of minimal (V) discrepancies of ciprofloxacin against E. faecalis, the numerous minor (III) and slight (IV) discrepancies of chloramphenicol against E. faecium, and the not insignificant number of very major (I) and minor (III) discrepancies observed with meropenem against isolates of E. faecalis necessitated proposals for new disk intermediate susceptibility criteria.     

Connexin expression and intercellular communication in two- and three-dimensional in vitro cultures of human bladder carcinoma

The identification of gap-junctional proteins (connexins) and the preparation of related antibodies provides new tools to study patterns of intercellular communication in tumors. Focusing on the biology of human bladder carcinoma, we compared the expression of gap-junctional proteins (connexins Cx26, Cx32, and Cx43) with a dye-coupling assay for gap-junctional intercellular communication in three cell lines with different urothelial differentiation. The cell lines HCV-29, RT4, and J82 were initially grown as monolayers of different ages. Connexin expression was found mostly positive over the time of culture and found constantly negative only in J82 cells for Cx26 and HCV-29 cells for Cx32. In HCV-29 cells, Cx26 increased in positivity over the time of culture. Western blotting with the antibodies confirmed the findings. Comparisons of dye transfer using Lucifer Yellow showed an increase of coupling in the normal urothelial cell line HCV-29 in contrast to a decrease of coupling in the tumor cell lines. Data were extended by multicellular spheroid (MCS) co-cultures with the stromal fibroblast line N1. In three-dimensional cultures as MCSs, Cx26 was increased in proximity of RT4 tumor cells to fibroblasts, and positivity was maintained in J82 cells. E-cadherin expression in cell lines showed no change in dependence of growth state. The data suggest that Cx26 plays a role in negative growth control or differentiation of urothelial cells. Preliminary comparative data on normal and neoplastic urothelium show all three connexins in normal urothelium, in contrast to varying amounts of Cx43 and low amounts of Cx32 in tumors and evident loss of Cx26 in low-grade tumors. Discrepancies between monolayer and MCS cultures are most likely due to higher differentiation in MCSs, and the continuation of systematic work with heterologous MCSs is indicated for more information on the role of gap-junctional proteins in human tumors.     

Cure of hyperparathyroidism in pregnancy by sternotomy and removal of a mediastinal parathyroid adenoma

Primary hyperparathyroidism is rarely reported during pregnancy but can cause significant maternal and neonatal morbidity. We report a case of hyperparathyroidism during pregnancy requiring a median sternotomy for resection of a mediastinal parathyroid adenoma. Surgery resulted in normalisation of serum calcium, resolution of symptoms, and prevented neonatal hypocalcaemia.     

Transplacental uptake of glucose is decreased in embryonic lethal connexin26-deficient mice

Mice that harbor a targeted homozygous defect in the gene coding for the gap junctional protein connexin26 died in utero during the transient phase from early to midgestation. From day 10 post coitum onwards, development of homozygous embryos was retarded, which led to death around day 11 post coitum. Except for growth retardation, no gross morphological alterations were detected between homozygous connexin26-defective embryos and wild-type littermates. At day 9 postcoitum, when chorioallantoic placenta started to function, connexin26 was weakly expressed in the yolk sac epithelium, between syncytiotrophoblasts I and II in the labyrinth region of the placenta, and in the skin of the embryo. At day 10 post coitum, expression of connexin26 in the placenta was much stronger than at the other locations. To analyze involvement of connexin26 in the placental transfer of nutrients, we have measured embryonic uptake of the nonmetabolizable glucose analogue 3-O-[14C]methylglucose, injected into the maternal tail vein. At day 10 post coitum, viable, homozygous connexin26-defective embryos accumulated only approximately 40% of the radioactivity measured in wild-type and heterozygous littermates of the same size. We conclude that the uptake of glucose, and presumably other nutrients as well, from maternal blood into connexin26-deficient mouse embryos was severely impaired and apparently not sufficient to support the rapid organogenesis during midgestation. Our results suggest that connexin26 gap junction channels likely fulfill an essential role in the transfer of maternal nutrients and embryonic waste products between syncytiotrophoblast I and II in the labyrinth layer of the mouse placenta.     

A One‐Pot “Triple‐C” Multicyclization Methodology for the Synthesis of Highly Constrained Isomerically Pure Tetracyclic Peptides

A broadly applicable one‐pot methodology for the facile transformation of linear peptides into tetracyclic peptides through a chemoenzymatic peptide synthesis/chemical ligation of peptides onto scaffolds/copper(I)‐catalyzed reaction (CEPS/CLIPS/CuAAC; “triple‐C”) locking methodology is reported. Linear peptides with varying lengths (≥14 amino acids), comprising two cysteines and two azidohomoalanines (Aha), were efficiently cyclized head‐to‐tail by using the peptiligase variant omniligase‐1 (CEPS). Subsequent ligation–cyclization with tetravalent (T4_1/2) scaffolds containing two bromomethyl groups (CLIPS) and two alkyne functionalities (CuAAC) yielded isomerically pure tetracyclic peptides. Sixteen different functional tetracycles, derived from bicyclic inhibitors against urokinase plasminogen activator (uPA) and coagulation factor XIIa (FXIIa), were successfully synthesized and their bioactivities evaluated. Two of these (FF‐T4_1/2) exhibited increased inhibitory activity against FXIIa, compared with a bicyclic control peptide. The corresponding hetero‐bifunctional variants (UF/FU‐T4_1/2), with a single copy of each inhibitory sequence, exhibited micromolar activities against both uPA and FXIIa; thus illustrating the potential of the “bifunctional tetracyclic peptide” inhibitor concept.     

Directed Divergent Evolution of a Thermostable D‐Tagatose Epimerase towards Improved Activity for Two Hexose Substrates

Functional promiscuity of enzymes can often be harnessed as the starting point for the directed evolution of novel biocatalysts. Here we describe the divergent morphing of an engineered thermostable variant (Var8) of a promiscuous D ‐tagatose epimerase (DTE) into two efficient catalysts for the C3 epimerization of D ‐fructose to D ‐psicose and of L ‐sorbose to L ‐tagatose. Iterative single‐site randomization and screening of 48 residues in the first and second shells around the substrate‐binding site of Var8 yielded the eight‐site mutant IDF8 (ninefold improved k_cat for the epimerization of D ‐fructose) and the six‐site mutant ILS6 (14‐fold improved epimerization of L ‐sorbose), compared to Var8. Structure analysis of IDF8 revealed a charged patch at the entrance of its active site; this presumably facilitates entry of the polar substrate. The improvement in catalytic activity of variant ILS6 is thought to relate to subtle changes in the hydration of the bound substrate. The structures can now be used to select additional sites for further directed evolution of the ketohexose epimerase.     

Model studies for wheat sourdough systems using gluten,lactate buffer and sodium chloride

Model studies were conducted in order to study the influence of acid and NaCl, as occurring in wheat sourdough bread, on fundamental rheological properties of wheat gluten. Gluten was divided into pieces and subjected to a swelling period in lactate buffer of pH 3.9, with or without added NaCl (3 g/100 ml). The respective controls were unbuffered NaCl solution and pure water. The microstructure of the gluten pieces was studied by laser-scanning confocal microscopy and the recombined pieces were examined using fundamental rheology. The combination of buffer (pH 3.9) and NaCl in comparison to unbuffered NaCl solution caused a denser, but partially dissolved fibrillar microstructure. Further to this, swelling of the gluten was reduced (62.0% versus 65.9% moisture) and an increase in firmness and elasticity was observed: in comparison with unbuffered NaCl solution, the absolute value of the complex dynamic modulus (|G*|) was higher, while the phase angle was lower in dynamic oscillatory measurements at 30 °C and dynamic temperature sweeps (30–95 °C), while in creep tests at 30 °C and 95 °C strain values were lower and relative recovery higher. In contrast, pH 3.9 buffer without added NaCl caused softer rheological behaviour than water and a film-like microstructure.     

The Influence of Dietary Habits and Meat Consumption on Plasma 3‐Methylhistidine—A Potential Marker for Muscle Protein Turnover

1 Scope

3‐Methylhistidine (3‐MH) as a potential biomarker for muscle protein turnover is influenced by meat intake but data on the impact of meat on plasma 3‐MH are scarce. We determined the association of plasma 3‐MH, 1‐methylhistidine (1‐MH), and creatinine with dietary habits and assessed the impact of a single white meat intervention during a meat‐free period.

2 Methods and results

Plasma 3‐MH, 1‐MH, and creatinine concentrations of healthy young omnivores (n = 19) and vegetarians (n = 16) were analyzed together with data on anthropometry, body composition, grip strength, and nutrition. After baseline measurements omnivores adhered to a meat‐free diet for 6 days and received a defined administration of chicken breast on day four. At baseline, omnivores had higher plasma 3‐MH and 1‐MH concentrations than vegetarians. White meat administration led to a slight increase in plasma 3‐MH in omnivores. The elevated 3‐MH concentrations significantly declined within 24 h after white meat intake.

3 Conclusion

1‐MH concentrations in plasma seem to be suitable to display (white) meat consumption and its influence on 3‐MH plasma concentration. 3‐MH in plasma may be used as a biomarker for muscle protein turnover if subjects have not consumed meat in the previous 24 h.