A Pilot Study of Femoropopliteal Artery Revascularisation with a Low Dose Paclitaxel Coated Balloon: Is Predilatation Necessary?

OBJECTIVE/BACKGROUND: The objective was to compare 2 year outcomes in patients treated with or without predilatation prior to drug coated balloon (DCB) angioplasty for symptomatic femoropopliteal lesions. METHODS: This prospective multicentre pilot study was conducted at three sites in Germany. It compared claudicants undergoing predilatation with a bare percutaneous transluminal angioplasty (PTA) balloon before DCB (predilatation group) with patients undergoing direct DCB (direct DCB group). Patients were followed for 2 years. Outcomes included late lumen loss at 6 months, and ankle brachial index (ABI), major adverse events, and primary patency at 2 years. A Clinical Events Committee and core laboratories analysed adverse events and angiographic/duplex images, respectively. RESULTS: Between December 2011 and November 2012, 50 patients were enrolled to the predilatation group (12% total occlusions) and 28 to the direct DCB group (5% total occlusions). Follow-up compliance at the 2 year visit was 88% (n = 44) and 86% (n = 24), respectively. Late lumen loss at 6 months was lower in the direct DCB group (0.03 ± 0.68 mm vs. 0.54 ± 0.97 mm; p = .01). Major adverse events over 2 years occurred in seven (15%) patients who underwent predilatation and in five (19%) after direct DCB. Mean ABI at 2 years was 0.94 ± 0.15 after predilatation and 1.0 ± 0.12 after direct DCB. Over 2 years, primary patency (80.3% vs. 78.2%; p = .55) was not statistically different between the groups. After propensity score adjustments, 2 year findings remained unchanged. CONCLUSION: Paclitaxel coated PTA, with or without bare predilatation, is effective over 2 years in symptomatic patients with femoropopliteal stenotic lesions. Adequately powered randomised controlled comparisons are required to confirm these preliminary results.

Effects of lactation on sodium intake in Fischer-344 and Long-Evans rats.

Rats of most strains are attracted to salt in low concentrations, and this attraction is increased by pregnancy and lactation. Nonreproducing Fischer-344 (F344) rats are unusual in that they avoid saline at all concentrations, raising the question of whether lactation alters their sodium appetite. Therefore, lactating and cycling Long-Evans and F344 rats were compared in their relative consumption of water and several concentrations of saline. We found that Long-Evans rats preferred saline, but F344 rats preferred plain water. In comparison with cycling rats, lactating Long-Evans rats markedly increased saline intake whereas F344 rats exhibited only a modest increase. Lactating rats of both strains increased total fluid intake, but this increase was primarily from water in F344 rats and from saline in Long-Evans rats. It was concluded that the aversion to saline characteristic of nonreproducing adult F344 rats continues during lactation. Their aversion to sodium in pup urine may account for the low levels of maternal anogenital licking observed in the F344 strain.

A fluorescence spectroscopy study of the calpactin I complex and its subunits p11 and p36: calcium-dependent conformation changes.

A fluorescence study of the calpactin I complex, a heterotetramer composed of two molecules of p36 and two molecules of p11, and its subunits, was performed to clarify their conformation. The analysis of the fluorescence characteristics of the single Trp of p36, in the absence of Ca(2+), shows that: (i) in the complex, Trp is buried within the protein matrix and subjected to static quenching from nearby groups; (ii) for p36 the results are similar, but Trp seems even more shielded than in the complex. Adding Ca(2+) to the calpactin I complex, or to p36, shifts the Trp emission maximum wavelengths, and increases the quantum yields which reflect a conformational change, burying the Trp in a more hydrophobic environment. In the presence and even in the absence of Ca(2+), the binding of phosphatidylserine liposomes induces a conformational change, detected by fluorescence measurements. The Ca(2+) dissociation constants, as determined by fluorescence titrations, are similar for the complex and p36 (KD approximately 0.5 x 10(-3) M). The affinity is enhanced a 1000-times in the presence of negatively charged phospholipids. In p11, both Try residues are located in a hydrophobic environment and the protein fluorescence does not change upon Ca(2+) addition.

Influence of the guanine nucleotide phosphorylation state and of Mg2+ ions on the interaction of vinzolidine/tubulin 6 S: a fluorescence quenching study.

The binding of the new vincaalkaloid vinzolidine to tubulin 6 S was investigated by using fluorescence quenching methods. The value of the apparent equilibrium binding constant was found to depend on the phosphorylation state of the guanine nucleotide bound to the tubulin exchangeable site (E-site), with Ka values of 4.9 X 10(4) and 8.19 X 10(4) M-1 for GTP- and GDP-tubulin, respectively. The effect of Mg2+ ions on this binding was more important on GTP-tubulin than on GDP-tubulin, and might be related to the existence of Mg2+ site(s) independent of the nucleotide.

Spectral properties of fluorescent derivatives of the oligomycin sensitivity conferring protein and analysis of their interaction with the F1 and F0 sectors of the mitochondrial ATPase complex.

In order to study the kinetics and the nature of the interactions between the oligomycin sensitivity conferring protein (OSCP) and the F0 and F1 sectors of the mitochondrial ATPase complex, fluorescent derivatives of OSCP, which are fully biologically active, have been prepared by reaction of OSCP with the following fluorescent thiol reagents: 6-acryloyl-2-(dimethylamino)naphthalene (acrylodan), 2-(4-maleimidylanilino)naphthalene-6-sulfonic acid (Mal-ANS), N-(1-pyrenyl)maleimide (Mal-pyrene), 7-(diethylamino)-3-(4-maleimidylphenyl)-4-methylcoumarin (Mal-coumarin), and fluorescein 5-maleimide (Mal-fluorescein). The preparation of these derivatives was based on the previous finding that the single cysteinyl residue of OSCP, Cys 118, can be covalently modified by alkylating reagents without loss of biological activity [Dupuis, A., Issartel, J. P., Lunardi, J., Satre, M., & Vignais, P. V. (1985) Biochemistry 24, 728-733]. For all fluorescent probes used, except Mal-pyrene and Mal-fluorescein, the emission spectra of conjugated OSCP were blue-shifted relative to those of the corresponding mercaptoethanol adducts, indicating that the fluorophores attached to Cys 118 were located in a hydrophobic pocket. These results were consistent with the high quantum yields and the increased fluorescence lifetimes of conjugated OSCP compared to mercaptoethanol adducts in aqueous buffer. They also fit with quenching data obtained with potassium iodide which showed that the fluorophore is shielded from the aqueous medium when it is attached to Cys 118 of OSCP. Especially noticeable was the wide half-width of the OSCP-acrylodan emission peak compared to that of mercaptoethanol-acrylodan.(ABSTRACT TRUNCATED AT 250 WORDS)

Conformational analysis of cholecystokinin CCK26-33 and related fragments by 1H NMR spectroscopy, fluorescence-transfer measurements, and calculations.

The conformational behavior of CCK7, Tyr-Met-Gly-Trp-Met-Asp-Phe-NH2, and CCK8, Asp-Tyr-Met-Gly-Trp-Met-Asp-Phe-NH2, in their sulfated and unsulfated forms, was studied both by 1H NMR spectroscopy in dimethyl-d6 sulfoxide and water and by fluorescence-transfer measurements at pH 7. In neutral conditions, both experimental methods show that these peptides exist preferentially in folded forms with beta and gamma turns around the sequence Gly-Trp-Met-Asp and Met-Asp-Phe-NH2, respectively. The presence of stable folded conformations is supported by through-space effects during the titration of the ionizable groups and by the weak temperature dependency of some amide protons not only in dimethyl sulfoxide but also in water. The folding of the C-terminal part, already shown in CCK5, seems to be a common conformational characteristic in CCK peptides. The N-terminal part of CCK8 presents an equilibrium between beta and gamma turns, whereas this part of the peptide is more flexible in CCK7. The low quantum yield of Tyr and the large mean distance (R = 15 A) between Tyr and Trp, determined by fluorescence-transfer measurements, support the occurrence of folded conformations pushing the aromatic rings far from each other. Interestingly, the introduction of the sulfate group enhances the folding tendency even in aqueous medium. The larger amide temperature dependency and the decrease in the R distance at acidic pH suggest that an intramolecular ionic interaction involving the N-terminal amino group and the beta-carboxyl groups of Asp32 stabilize the folded forms. Metropolis calculations performed on CCK8 support the existence of stable folded conformations closely related to those deduced from experimental data.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of cholera toxin and pertussis toxin on opioid tolerance and dependence in the guinea-pig myenteric plexus.

Opioid tolerance and dependence are characterized in terms of inhibitory and excitatory neuronal mechanisms. Bacterial toxins were used to investigate these phenomena in the isolated guinea-pig ileum (GPI) and mouse vas deferens (MVD). Cholera toxin (CT) and pertussis toxin (islet activating protein; IAP) have been demonstrated to selectively impair the function of either the stimulatory or inhibitory nucleotide regulatory protein, mediating the signal transmission to the adenylate cyclase. It has been found that CT failed to affect electrically evoked twitch tension in naive and tolerant GPI and MVD. Twitch tension evoked by excitatory drugs, e.g. neurotensin, was attenuated in the GPI, but not in the MVD. CT did not interfere with the actions of opioids or nonopioids in either the GPI or the MVD. Similarly, IAP failed to affect electrically evoked twitch tension. It virtually lacked an effect on the inhibitory action of opioids and nonopioids on electrically stimulated naive and tolerant GPI and MVD. On the other hand, both CT and IAP dose-dependently attenuated the naloxone precipitated withdrawal contracture in the GPI rendered opioid-dependent. These findings may suggest that opioid receptors at nerve terminals of the GPI and the MVD are not linked to adenylate cyclase, and those receptors relate to the state of tolerance. In contrast, opioid receptors located at nerve somata may be linked to adenylate cyclase. Those receptors may be associated with the development of dependence. The experiments with CT suggest a synaptic excitatory input of neighboring nerve elements necessary for the generation of a withdrawal contracture.

Opioid dependence prevents the action of pertussis toxin in the guinea-pig myenteric plexus.

The longitudinal muscle-myenteric plexus preparation of the guinea-pig ileum has been employed for the study of the effect of pertussis toxin (IAP) on opioid dependence. Guinea-pigs were treated with IAP (120 micrograms/kg, i.p.) either prior to chronic administration of an opioid or after opioid dependence had been established. The isolated preparations were tested in vitro for dependence; that is, the naloxone-precipitated withdrawal contracture. Naloxone almost failed to evoke a sign of dependence in preparations treated with IAP prior to chronic exposure to an opioid. In contrast, IAP failed to affect the withdrawal contracture when applied to an animal after dependence has been established. It is concluded that the Ni-unit, the substrate for IAP, plays a critical function in the development of dependence. The continuous activation of the opioid receptor associated with the development of dependence may induce changes in Ni which in turn prevent the interaction of IAP with its substrate.

Experimental systemic toxicology of 4'-epidoxorubicin, a new, less cardiotoxic anthracycline antitumor agent.

The acute and chronic iv toxicity of 4'-epidoxorubicin, a new antitumor anthracycline antibiotic, was compared with doxorubicin. The LD50 of 4'-epidoxorubicin was 16.07 mg/kg in mice, 14.27 mg/kg in rats, and about 2 mg/kg in dogs; the LD50 of doxorubicin was 11.98 mg/kg in mice, 10.51 mg/kg in rats, and about 2.5 mg/kg in dogs. Rats and dogs were also dosed iv for 91 days (3 injections/week) with 4'-epidoxorubicin or doxorubicin at doses of 0.128, 0.32, and 0.8 mg/kg to rats and 0.064, 0.16, and 0.4 mg/kg to dogs. A comprehensive toxicological evaluation of the animals was carried out before, throughout, and at the end of the study. High-dose 4'-epidoxorubicin induced toxic clinical signs in dogs, and in both species caused loss of body weight, antiproliferative effects on blood-forming organs and testes, and degenerative lesions in kidneys and heart. The cardiac damage was moderate in rats and very mild in dogs; only three male rats died at this dose. The medium dose induced less pronounced changes and no heart lesions and the low dose was practically nontoxic. Doxorubicin showed similar antiproliferative activity, but more evident toxic effects, especially on the heart; many rats given the high dose died and some at the medium dose showed initial cardiac lesions. Thus 4'-epidoxorubicin appeared less toxic than doxorubicin; in particular cardiac damage was much less evident in animals chronically injected with the new drug.

Intrinsic fluorescence of a non-myelin apoproteolipid and evidence for the existence of conformational flexibility.

An extremely hydrophobic protein (Mr = 16000), which in its native form is only soluble in organic solvents and which differs from the myelin proteolipid (Mr = 24000), was purified to homogeneity. Intrinsic fluorescence studies on this apoproteolipid have revealed a large conformational flexibility. In the water-soluble form the emitting residues appear to be buried in a hydrophobic core while in organic solvents they are exposed to the external medium. Structural changes depending on the organic solvent are also observed. The emission characteristics of reconstituted proteoliposomes may be due to the formation of a membrane-linked complex between several proteolipid monomers.

Long-term observation of young cystinuric patients under ascorbic acid therapy.

Long-term results of ascorbic acid monotherapy in four young patients with cystine stone complaints are reported. The therapeutic program is based on the descriptions by Asper and Schmucki, who publicized the method for the first time in 1979. Clinical course observations, renal function checks as well as the excretion of cystine, uric acid and oxalic acid during ascorbic acid therapy of our patient pool will be discussed. Clinical observations made to date indicate that ascorbic acid therapy is a practical, inexpensive prophylaxis for cystine stone patients that is virtually free of side effects.