A high-resolution NMR study (1H, 13C, 31P) of the interaction of paramagnetic ions with phospholipids in aqueous dispersions

In the surgical treatment of hernias in children, many differences of opinion still exist, especially in the selection of the type of procedure to be used. The authors have applied the Ferguson Method in the repair of Inguinal hernias as a method of choice. Along with a high resection of Processus Vaginalis of the peritoneum, the anterior wall of Canalis Inguinalis is reinforced without transposing the Funiculus Spermaticus. To evaluate the success of this method, the authors made a late postoperative analysis. Examinations were made on 106 children of various ages who had unilateral inguinal hernia repairs one to two years prior. The unoperated contralateral inguinal canal served as a control by which eventual changes of testis could be evaluated. Clinical examinations involved determination of whether or not a recidive hernia exits, as well as the condition, size and consistency of the testis. Accordingly, poor postoperative results using the Ferguson method were less than one percent.     

2H NMR studies of a myristoylated peptide in neutral and acidic phospholipid bicelles

Deuterium NMR spectra of Myr-d27-GNAAAAKKGSEQES (Cat14), the N-terminal 14-residue peptide from the catalytic subunit of cAMP-dependent protein kinase A (PKA), illustrate how magnetically aligned neutral and acidic phospholipid bicelles can be used to characterize the ordering and mode of binding of peptides to membranes. Since Cat14 is electrically neutral, the major interaction responsible for binding is the insertion of the myristoyl group into the hydrophobic core of the bilayer. The inclusion of 25% phosphatidylserine or phosphatidylglycerol into phosphatidylcholine bicelles results in a moderate increase in the ordering of the peptide relative to the bicelle normal, presumably because of favorable electrostatic interactions between the phospholipid headgroups and the two lysines in positions 7 and 8. Successful preparation of acidic bicelles was achieved by careful adjustment of lipid composition, pH and ionic strength.     

Effect of sugars on headgroup mobility in freeze-dried dipalmitoylphosphatidylcholine bilayers: solid-state 31P NMR and FTIR studies

The effect of the carbohydrates trehalose, glucose, and hydroxyethyl starch (HES) on the motional properties of the phosphate headgroup of freeze-dried dipalmitoylphosphatidylcholine (DPPC) liposomes was studied by means of 31P NMR, Fourier transform infrared spectroscopy (FTIR), and differential scanning calorimetry (DSC). The results show that trehalose, which is a strong glass former (Tg = 115 degreesC), elevates the onset of the lipid headgroup rotations and preserves some rotational mobility of the phosphate headgroups after cooling from the liquid-crystalline state. Glucose (Tg = 30 degreesC), a very effective depressant of the phase transition temperature of freeze-dried DPPC, markedly elevates the initiation of the temperature of headgroup rotations. On the other hand, the monosaccharide does not preserve the headgroup disordering when cooled from the liquid-crystalline state. These effects are consistent with formation of hydrogen bonds between the OH groups of the sugar and the polar headgroups of DPPC. They show, however, that hydrogen bonding is not sufficient for preservation of the dynamic properties of freeze-dried DPPC. HES, although a very good glass former (Tg > 110 degreesC), does not depress the phase transition temperature and affects only slightly the rotational properties of freeze-dried DPPC. This lack of effect of HES is associated with the absence of direct interactions with the lipid phosphates, as evidenced by the FTIR results. These data show that vitrification of the additive is not sufficient to affect the dynamic properties of dried DPPC.     

Effect of 6-aminonicotinamide on the pentose phosphate pathway: 31P NMR and tumor growth delay studies

6-aminonicotinamide (6AN) has been shown to enhance radiosensitivity in vitro, although previous in vivo studies failed to show an effect. 31P NMR spectra were obtained by using a one-dimensional chemical shift imaging technique on a first generation transplant of the CD8FI spontaneous mammary carcinoma tumor model. Spectra were obtained both before and 10 h after treatment with 6AN (20 mg/kg). Changes in pH, nucleoside triphosphate/inorganic phosphate, and phosphocreatine/ inorganic phosphate measured at 10 h post-6AN were not significant. A new peak was detected 10 h post-6AN, which was assigned to 6-phosphogluconate (6PG), indicating inhibition of the pentose phosphate pathway (PPP). Based on the spectral data demonstrating inhibition of the PPP at 10 h post-6AN, tumor-bearing mice were irradiated (15 Gy x 3 fractions) on Days 1, 10 or 11, and 21 10 h after administration of 6-aminonicotinamide (20 mg/kg). Tumor-bearing mice receiving 6AN alone (20 mg/kg x 3), radiation alone (15 Gy x 3), or saline were also studied. Tumor growth delay studies indicated that 6AN alone induced a small but significant tumor growth delay (4.3 +/- 0.8 days). Radiation alone induced a tumor growth delay of 34.5 +/- 2.7 days. Treatment with 6AN followed by radiation induced a tumor growth delay of 57.0 +/- 3.8 days. This was significantly greater than the TGD values for treatment with 6AN alone or radiation (P < 0.01). No complete regressions were noted after treatment with 6AN or radiation alone. Concomitant therapy with 6AN plus radiation yielded 6/28 complete regressions (21%), which was significantly greater than radiation (P < 0.05) or 6AN alone (P < 0.01) on this mammary carcinoma.     

Effects of divalent cations, protons and calmidazolium at the rat P2X7 receptor

The P2X7 receptor is a uniquely bifunctional molecule through which ATP can open a small cationic channel typical of ionotropic receptors and also induce a large pore permeable to high molecular weight molecules (> 600 Da). Activation of this large pore can lead to cell lysis within 1-2 min. We asked whether pharmacological differences existed between the cationic channel and the cell permeabilizing pore by measuring whole-cell currents and uptake of a propidium dye (YO-PRO; Mw 629) in HEK293 cells stably expressing the rat P2X7 receptor, and comparing the actions of divalent cations and protons in these two assays. Currents in response to 2'-3'-(O)-(4-benzoyl benzoyl) ATP (BzATP, 30 microM) were inhibited by extracellular calcium, magnesium, zinc, copper and protons with half-maximal inhibitory concentrations (IC50) of 2.9 mM, 0.5 mM, 11 microM, 0.5 microM and 0.4 microM, respectively. The inhibition was voltage independent in each case. YO-PRO uptake induced by BzATP was also inhibited with similar IC50 values. The rank order of potency of a range of divalents was Cu2+ > Cd2+ = Zn2+ > Ni2+ > Mg2+ = Co2+ > Mn2+ > Ca2+ = Ba2+ > Sr2+. These results suggest that these divalent cations and protons all act primarily as allosteric modulators to alter the affinity of ATP binding to the P2X7 receptor. In contrast, extracellular (but not intracellular) calmidazolium inhibited the BzATP-evoked current by up to 90% (IC50 = 15 nM) but had no effect on YO-PRO uptake. Thus, calmidazolium can block activation of the ionic channel but this does not prevent the formation of the large permeabilizing pore.     

Controllable graded cerebral ischaemia in the gerbil: studies of cerebral blood flow and energy metabolism by hydrogen clearance and 31P NMR spectroscopy

Patent ductus arteriosus is an uncommon anomaly in adult patients. Surgical closure of patent ductus arteriosus in this age group presents difficult problems to the surgeon. We report our experience of 21 adult patients (19-62 years of age, mean 40 years) who underwent closure of the ductus by transfemoral implantation of a Rashkind double umbrella device. The patients came to light because of atrial fibrillation, congestive heart failure, residual flow after surgical ligation of the duct or because of incidental diagnosis made during physical examination or chest X-ray. In ten patients the pulmonary arterial pressure was normal (systolic pressure < 30 mmHg), in eleven it was elevated (systolic pressure from 30 to 100 mmHg, mean 50 mmHg). In seven patients the duct was clearly calcified and the size of the duct varied from 3 to 9 mm (mean 4.3 mm). In 16 patients the ductus resulted perfectly closed after implantation of the first double umbrella device, two patients had minimal residual aortopulmonary flow, whereas in three patients the residual shunt was significant; two of these also developed haemolysis and went to surgery, in the latter the shunt was completely abolished after implantation of a second 17-mm device 16 months later. In conclusion transcatheter closure of patent ductus arteriosus in adults is feasible, even in the presence of calcifications and/or pulmonary hypertension; taking into account the significant surgical risk, PDA umbrella closure should be considered the first choice procedure in this group of patients.     

Detection and quantitation of phosphorus metabolites in crude tissue extracts by 1H and 31P NMR: use of gradient assisted 1H-31P HMQC experiments, with selective pulses, for the assignment of less abundant metabolites

The lungs and blood of Yusho patients and healthy Japanese people have already been contaminated with methylsulphonyl polychlorinated biphenyls (MSF-PCBs) at relatively high concentration. Therefore, we should give due attention to their biological and toxicological effects to man. In this study, in order to mainly evaluate non-S-dependent genotoxicity of five MSF-PCB congeners, namely, 3-MSF-4, 5, 3', 4'-tetrachlorobiphenyl (TCB), 3-MSF-4, 5, 2', 3'-TCB, 3-MSF-2, 5, 2', 4', 5'-pentachlorobiphenyl (PenCB), 4-MSF-2, 5, 2', 3', 4'-PenCB and 4-MSF-2, 5, 2', 3', 5', 6'-hexachlorobiphenyl (HCB). We have examined their effects on the induction of micronucleated cells, which has been frequently used to estimate the dose of ionizing radiation and truly radiomimetic, non-S-dependent, clastogens, in cultured human lymphocytes in the absence or presence of 2, 3, 4, 7, 8-pentachlorodibenzofuran (PenCDF), 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD) or 3, 4, 5, 3', 4'-pentachlorobiphenyl (Co-PenCB). The following results were obtained. 1) 4 x 10(-5) M7, 8-benzoflavone (ANF) significantly enhanced the frequency of micronucleated cells and all of the five MSF-PCB congeners failed to induce the formation of micronucleated cells at doses of 5.2 to 9.6 ppm, which were about 35,000 times higher than the concentrations in the lungs and adipose tissue of healthy Japanese people.(ABSTRACT TRUNCATED AT 250 WORDS)     

Direct administration and utilization of [1-13C]glucose by fetal brain and liver tissues under normal and ischemic conditions: 1H, 31P, and 13C NMR studies

The acute and delayed effects of anoxia on synaptic transmission and long-term potentiation (LTP) were examined in the CA1 region of rat hippocampal slices. Oxygen deprivation for 20 minutes completely but reversibly depressed excitatory postsynaptic potentials mediated by both N-methyl-D-aspartate receptors (NMDAR) and non-NMDAR. Although LTP was reliably produced by a single tetanus delivered 30 minutes after reoxygenation, LTP could not be induced when a tetanus was delivered 70 to 100 minutes after reoxygenation. A tetanus delivered 100 minutes after reoxygenation produced lasting synaptic enhancement when 100 mumol/L D,L-amino-phosphonovaleric acid (APV), a competitive NMDAR antagonist, was administered during the period of oxygen deprivation. The delayed effects of oxygen deprivation were not blocked when APV was administered after oxygen deprivation. Similarly, the delayed effects on LTP induction were overcome by inhibitors of nitric oxide synthase when the nitric oxide synthase inhibitors were administered during anoxia, but not when administered after oxygen deprivation. These results suggest that untimely activation of NMDAR and nitric oxide release during anoxia produce delayed inhibition of LTP induction and may be involved in the memory defects that occur subsequent to cerebral hypoxia.     

Nucleoside 3'-phosphotriesters as key intermediates for the oligoribonucleotide synthesis. IV. New method for removal of 2,2,2-trichloroethyl group and 31P NMR as a new tool for analysis of deblocking of internucleotide phosphate protecting groups

Zinc/acetylacetone/pyridine treatment has been designed as a very efficient method for removal of 2,2,2,-trichloroethyl group from phosphoesters. Internucleotide and terminal 2,2,2-trichloroethylphosphotriesters were transformed to corresponding diesters quantitatively. Much less reactive 2,2,2-trichloroethylphosphodiesters produced monoesters with ca. 90% yield. 31P NMR spectroscopy has been proposed as a new tool for analysis of removal of internucleotide phosphate protecting groups-a crucial step in oligonucleotides synthesis via phosphotriester approach.