Edith Hantz

Aminoglycoside and glycopeptide renal toxicity in intensive care patients studied by proton magnetic resonance spectroscopy of urine.

ObjectiveAminoglycoside and glycopeptide antibiotics are responsible for renal toxicity. In most cases, the nephrotoxicity is limited to a reversible tubular injury, but an acute and sustained renal failure may occur. The aim of our study was to explore the renal function of patients given these antimicrobial agents with proton magnetic resonance spectroscopy of urine. This technique is able to detect, in urine samples, a wide range of metabolites reflecting renal tubular function. The variables assessed by magnetic resonance spectroscopy were compared with the routine markers of renal function: creatinine, urea, and 24-hr urine volume. DesignProspective clinical study. SettingIntensive care unit. PatientsAll patients in an intensive care unit receiving an aminoglycoside and/or a glycopeptide were included in the study if they presented with signs of renal dysfunction. All experiments were performed on urine samples collected for the routine follow-up of these patients. InterventionProton spectra were acquired with water suppression, and the peak intensity of each metabolite was reported in relationship to the intensity of the creatinine peak. Measurements and Main ResultsThe ratio values obtained by magnetic resonance spectroscopy were compared with the values of creatininemia and blood urea obtained routinely by biochemistry and with the value of the 24-hr urine volume by logistic regression and general linear models. This statistical analysis showed that the ratio of dimethylamine to creatinine was highly correlated with creatininemia. ConclusionsDimethylamine is an osmolyte released from the medullar region of the kidney. Thus, our study demonstrated that nephrotoxicity from aminoglycosides and glycopeptides is not limited to proximal tubular toxicity but also may involve the medullar region (Henle loop and collecting duct) of the nephron.

The osmotic response of large unilamellar vesicles studied by quasielastic light scattering

Large unilamellar vesicles of two phosphatidylcholines, one saturated (DMPC) and the other unsaturated (DOPC), prepared by the reverse-phase evaporation method were studied using the quasielastic light scattering technique. The accurate sizing obtained by this technique showed an osmotic response for the two kinds of vesicles when the salinity of the external medium was diluted. The elastic moduli of lipid vesicles bilayers in the liquid phase were then estimated according to the elasticity theory of spherical shells taking into account salt leakage data known from the literature.

Solution structure of a truncated anti‐MUC1 DNA aptamer determined by mesoscale modeling and NMR

Mucin 1 is a well‐established target for the early diagnosis of epithelial cancers. The nucleotides of the S1.3/S2.2 DNA aptamer involved in binding to variable number tandem repeat mucin 1 peptides have been identified using footprinting experiments. The majority of these binding nucleotides are located in the 25‐nucleotide variable region of the total aptamer. Imino proton and 2D NMR spectra of truncated and total aptamers in supercooled water reveal common hydrogen‐bonding networks and point to a similar secondary structure for this 25‐mer sequence alone or embedded within the total aptamer. NMR titration experiments confirm that the TTT triloop structure is the primary binding site and show that the initial structure of the truncated aptamers is conserved upon interaction with variable number tandem repeat peptides. The thermal dependence of the NMR chemical shift data shows that the base‐paired nucleotides melt cooperatively at 47 ± 4 °C. The structure of the 25‐mer oligonucleotide was determined using a new combined mesoscale molecular modeling, molecular dynamics and NMR spectroscopy investigation. It contains three Watson–Crick pairs, three consecutive mispairs and four Watson–Crick pairs capped by a TTT triloop motif. The 3D model structures (PDB 2L5K) and biopolymer chain elasticity molecular models are consistent with both NMR and long unconstrained molecular dynamics (10 ns) in explicit water, respectively.

Magnetic resonance spectroscopy of cellular lipid extracts from sensitive, resistant and reverting K562 cells and flow cytometry for investigating the P-glycoprotein function in resistance reversion

The proton NMR spectra of K562 cells contain resonances of lipids. When these cells acquire multidrug resistance phenotype, the NMR lipid signals are modified and partially recovered when the resistance is reversed. The goals of the present study are to elucidate the mechanism of the resistance phenotype reversion and to investigate the possible origin of lipid signals detected in whole cells with proton NMR spectroscopy. Therefore, the K562 drug‐sensitive cell line, its adriamycin resistant counterpart and two reverting derivates, obtained by verapamil treatment and long term culture in drug‐free medium, were used in this study. The P‐glycoprotein (P‐gp) pump function was measured by flow cytometry and lipids were extracted to be analysed by proton and phosphorus spectroscopy. The phenotype reversion is due to the decrease of the P‐gp function and an increased entrance of anthracycline drug when compared with the resistant cells. The spectra obtained on extracts showed no modification of the fatty acid composition and of the ratio of total cholesterol to fatty acid content. A different phospholipid composition in sensitive and resistant cells was found, but the reversion of resistance did not produce a recovery of these lipids. Thus, the lipid NMR spectra of extracts could not explain the spectral modifications observed on whole cells, in relation to acquiring and reverting drug resistance. These results are in favour of a different lipid organization or of localization within the cell. Copyright

Solution structure of a purine rich hexaloop hairpin belonging to PGY/MDR1 mRNA and targeted by antisense oligonucleotides

A preferential target of antisense oligonucleotides directed against human PGY/MDR1 mRNA is a hairpin containing a stem with a G•U wobble pair, capped by the purine-rich 5′r(GGGAUG)3′ hexaloop. This hairpin is studied by multidimensional NMR and restrained molecular dynamics, with special emphasis on the conformation of south sugars and non-standard phosphate linkages evidenced in both the stem and the loop. The hairpin is found to be highly structured. The G•U wobble pair, a strong counterion binding site, displays structural particularities that are characteristic of this type of mismatch. The upper part of the stem undergoes distortions that optimize its interactions with the beginning of the loop. The loop adopts a new fold in which the single-stranded GGGA purine tract is structured in A-like conformation stacked in continuity of the stem and displays an extensive hydrogen bonding surface for recognition. The remarkable hairpin stability results from classical inter- and intra-strand interactions reinforced by numerous hydrogen bonds involving unusual backbone conformations and ribose 2′-hydroxyl groups. Overall, this work emphasizes numerous features that account for the well-ordered structure of the whole hairpin and highlights the loop properties that facilitate interaction with antisense oligonucleotides.

Study of emulsions of pharmaceutical interest by light scattering

Abstract Emulsions of light liquid paraffin oil in water have been studied by a light-scattering technique. These emulsions were stabilized with a mixture of simulsol 92 and simulsol 96 as surfactants in the critical HLB range. The influence of the dilution and of the physical parameters of the continuous phase on the translational-diffusion coefficient D a and that of the surfactant HLB, as a function of pH, salinity and temperature, have been established. The extrapolation to infinite dilution of D a leads to an accurate determination of the hydrodynamic radius of the droplets which is in the submicrometre range. On the other hand, the intensity data permitted an estimation of the gyration radius and provided insights into the surfactant layer thickness of the particles. The stability of the emulsions has been discussed in terms of the interaction potential barrier.

The effect of loperamide on the thermal behavior of dimyristoylphosphatidylcholine large unilamellar vesicles

The effect of loperamide, a drug belonging to the opiate family, on dimyristoyl phosphatidylcholine large unilamellar vesicles (DMPC LUV) was investigated by quasielastic light scattering (QLS) and Fourier transform infrared spectroscopy (FT-IR). Both techniques show that, in the presence of loperamide, DMPC LUV undergoes a two step transition in cooling: one step around the transition point of pure lipid vesicles, the other at a lower temperature. The temperature of the latter step transition is different for the head and tail regions of the drug-containing vesicles: FT-IR spectra demonstrate that the hydrophobic acyl chains transition starts at a temperature well above that of the interfacial region whereas the transition of the entire vesicle, explored by QLS, is broad and covers both temperature ranges. These transitions are thermally reversible in the FT-IR which measures local order but aggregation effects prevent the thermal reversibility of the QLS results. The nature of the drug-lipid interaction is also discussed.

Quasi-elastic light scattering study of salt induced conformational transitions of chromatin subunit

Abstract The translational diffusion coefficient D T of monodisperse solutions of 146 base pairs (bp) core particles was studied by the quasi-elastic light scattering technique. When the salinity was raised a change of D T from 1.9 × 10 −7 cm 2 s −1 to 3.2 × 10 −7 cm 2 s −1 was detected at about 2 m M NaCl, followed by a smooth decrease of D T beyond 0.6 M NaCl. The measurements of particle concentration and scattering vector effects on the D T showed that the influence of interactions between particles can be disregarded. The interaction between particles and counterions is also discussed and does not appear to be the origin of the actual changes in D T . These transitions of D T are hence related to changes of shape and size of the particles. It is shown that the single transition at low salinity corresponds to a conformational change while the variation of D T at high salinity can be interpreted by a destabilization of the edifice. In different regions of salinities, the observed values of D T can lead to reasonable hydrodynamic models.

Nuclear magnetic resonance metabolomics and human liver diseases: The principles and evidence associated with protein and carbohydrate metabolism (Review)

During the last decade, metabolomics has become widely used in the field of human diseases. Numerous studies have demonstrated that this is a powerful technique for improving the understanding, diagnosis and management of various types of liver disease, such as acute and chronic liver diseases, and liver transplantation. Nuclear magnetic resonance (NMR) spectroscopy is one of the two most commonly applied methods for metabolomics. The aim of the present review was to investigate the results from recent key publications focusing on aspects of protein and carbohydrate metabolism. The review includes existing procedures, which are currently used for NMR data acquisition and statistical analysis. In addition, notable results obtained by these studies on protein and carbohydrate metabolism concerning human liver diseases are presented.

Deuteration of Water Enables Self-Organization of Phospholipid-Based Reverse Micelles

Phospholipid-based reverse micelles are composed of branched cylinders. Their branching points are known to attract themselves and to slide along branches. The rate of this sliding is governed by the lifetime of H(D)-bonded water bridges between phospholipid molecules. This lifetime is increased when the water is deuterated. On condition that the water contains at least 40 D atoms%, water/dipalmitoylphosphatidylcholine (DPPC)/deuterated pyridine reverse micelles with the composition 1.1:1:250 (v/v) have been shown to self-organize into a liquid crystal in the 310-316 K temperature range. The mechanism of this self-organization is unraveled by following the FTIR and (1)H NMR spectra of more concentrated micelles upon heating. During the preparation of micelles, pyridine-(D(+))H(+) ions are formed. They give rise to hydron transfers, under the influence of the DPPC electric charges, evidenced by two broad FTIR absorptions above (BB1) and below (BB2) the nu(C-O) stretch. These hydron transfers occur along strong (D(+))H(+) bonds of pyridinium ions with pyridine (BB1) and DPPC C=O groups (BB2). The proton transfers at the interface of micelles, relayed in the continuous pyridine medium, create a tenuous link between separated micelles, thus facilitating their organization. Upon heating, DPPC heads shrink and DPPC chains expand to make wedge-shaped DPPC molecules. The micelles then change in shape: cylinders constrict and enclosed water drifts towards branching points, which swell. Branching points of neighboring micelles come into contact. Due to the deuteration of water these contacts are prolonged and H bonds are formed between DPPC molecules located in each branching point. Upon storage at 39 degrees C, these branching points fuse. The lateral diffusion of DPPC molecules becomes free, as evidenced by a narrowing of all (1)H NMR resonances. Upon further heating, reorganization into a liquid crystal occurs.