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61.
Upon topical administration, a high penetration rate of antifungal drug into the infected site is desirable to reduce treatment
length and systemic side effects which occur especially after a prolonged peroral administration. Thermogelling formulations
composed of poloxamer 407, medium chain triglycerides, isopropyl alcohol, dimethyl isosorbide, and water for topical application
were developed, and a lipophilic drug terbinafine HCl (TBF) was incorporated. Previously, a remarkable high permeation rate
of a hydrophilic drug 5-aminolevulinic acid from this vehicle was evident compared to different creams from German Pharmacopoeia.
By varying the composition of vehicle constituents, a broad range of consistencies and appearances was obtained. Up to 4%
TBF could be solubilized in the vehicle. TBF fluxes at steady state across human stratum corneum from these formulations were
higher than those from the German Pharmacopoeia Basiscreme Deutscher Arzneimittel Codex and a marketed product at similar
concentration of 1%. TBF fluxes increased along with a higher content of TBF in the formulation. The amount of TBF retained
in stratum corneum was higher compared to those from both standards of comparison (p < 0.01). The thermodynamic activity of TBF in the thermogelling formulation was lower compared to those in other formulations.
Therefore, the nature of the vehicle and its interaction with TBF are suggested to play a significant role in explaining higher
fluxes along with higher TBF content. Differential scanning calorimetry measurements revealed comparable T2 and T3 endothermic
shifts from all examined formulations suggesting equal influences to the skin lipids. 相似文献
62.
Pfefferle S Schöpf J Kögl M Friedel CC Müller MA Carbajo-Lozoya J Stellberger T von Dall'Armi E Herzog P Kallies S Niemeyer D Ditt V Kuri T Züst R Pumpor K Hilgenfeld R Schwarz F Zimmer R Steffen I Weber F Thiel V Herrler G Thiel HJ Schwegmann-Wessels C Pöhlmann S Haas J Drosten C von Brunn A 《PLoS pathogens》2011,7(10):e1002331
Coronaviruses (CoVs) are important human and animal pathogens that induce fatal respiratory, gastrointestinal and neurological disease. The outbreak of the severe acute respiratory syndrome (SARS) in 2002/2003 has demonstrated human vulnerability to (Coronavirus) CoV epidemics. Neither vaccines nor therapeutics are available against human and animal CoVs. Knowledge of host cell proteins that take part in pivotal virus-host interactions could define broad-spectrum antiviral targets. In this study, we used a systems biology approach employing a genome-wide yeast-two hybrid interaction screen to identify immunopilins (PPIA, PPIB, PPIH, PPIG, FKBP1A, FKBP1B) as interaction partners of the CoV non-structural protein 1 (Nsp1). These molecules modulate the Calcineurin/NFAT pathway that plays an important role in immune cell activation. Overexpression of NSP1 and infection with live SARS-CoV strongly increased signalling through the Calcineurin/NFAT pathway and enhanced the induction of interleukin 2, compatible with late-stage immunopathogenicity and long-term cytokine dysregulation as observed in severe SARS cases. Conversely, inhibition of cyclophilins by cyclosporine A (CspA) blocked the replication of CoVs of all genera, including SARS-CoV, human CoV-229E and -NL-63, feline CoV, as well as avian infectious bronchitis virus. Non-immunosuppressive derivatives of CspA might serve as broad-range CoV inhibitors applicable against emerging CoVs as well as ubiquitous pathogens of humans and livestock. 相似文献
63.
Sainlos M Tigaret C Poujol C Olivier NB Bard L Breillat C Thiolon K Choquet D Imperiali B 《Nature chemical biology》2011,7(2):81-91
The interactions of the AMPA receptor (AMPAR) auxiliary subunit Stargazin with PDZ domain-containing scaffold proteins such as PSD-95 are critical for the synaptic stabilization of AMPARs. To investigate these interactions, we have developed biomimetic competing ligands that are assembled from two Stargazin-derived PSD-95/DLG/ZO-1 (PDZ) domain-binding motifs using 'click' chemistry. Characterization of the ligands in vitro and in a cellular FRET-based model revealed an enhanced affinity for the multiple PDZ domains of PSD-95 compared to monovalent peptides. In cultured neurons, the divalent ligands competed with transmembrane AMPAR regulatory protein (TARP) for the intracellular membrane-associated guanylate kinase resulting in increased lateral diffusion and endocytosis of surface AMPARs, while showing strong inhibition of synaptic AMPAR currents. This provides evidence for a model in which the TARP-containing AMPARs are stabilized at the synapse by engaging in multivalent interactions. In light of the prevalence of PDZ domain clusters, these new biomimetic chemical tools could find broad application for acutely perturbing multivalent complexes. 相似文献
64.
65.
Crystel Bonnet M’hamed Grati Sandrine Marlin Jacqueline Levilliers Jean-Pierre Hardelin Marine Parodi Magali Niasme-Grare Diana Zelenika Marc Délépine Delphine Feldmann Laurence Jonard Aziz El-Amraoui Dominique Weil Bruno Delobel Christophe Vincent Hélène Dollfus Marie-Madeleine Eliot Albert David Catherine Calais Jacqueline Vigneron Bettina Montaut-Verient Dominique Bonneau Jacques Dubin Christel Thauvin Alain Duvillard Christine Francannet Thierry Mom Didier Lacombe Françoise Duriez Valérie Drouin-Garraud Marie-Françoise Thuillier-Obstoy Sabine Sigaudy Anne-Marie Frances Patrick Collignon Georges Challe Rémy Couderc Mark Lathrop José-Alain Sahel Jean Weissenbach Christine Petit Françoise Denoyelle 《Orphanet journal of rare diseases》2011,6(1):1-19
Background
Usher syndrome (USH) combines sensorineural deafness with blindness. It is inherited in an autosomal recessive mode. Early diagnosis is critical for adapted educational and patient management choices, and for genetic counseling. To date, nine causative genes have been identified for the three clinical subtypes (USH1, USH2 and USH3). Current diagnostic strategies make use of a genotyping microarray that is based on the previously reported mutations. The purpose of this study was to design a more accurate molecular diagnosis tool.Methods
We sequenced the 366 coding exons and flanking regions of the nine known USH genes, in 54 USH patients (27 USH1, 21 USH2 and 6 USH3).Results
Biallelic mutations were detected in 39 patients (72%) and monoallelic mutations in an additional 10 patients (18.5%). In addition to biallelic mutations in one of the USH genes, presumably pathogenic mutations in another USH gene were detected in seven patients (13%), and another patient carried monoallelic mutations in three different USH genes. Notably, none of the USH3 patients carried detectable mutations in the only known USH3 gene, whereas they all carried mutations in USH2 genes. Most importantly, the currently used microarray would have detected only 30 of the 81 different mutations that we found, of which 39 (48%) were novel.Conclusions
Based on these results, complete exon sequencing of the currently known USH genes stands as a definite improvement for molecular diagnosis of this disease, which is of utmost importance in the perspective of gene therapy. 相似文献66.
Tsai IJ Liu ZW Rayment J Norman C McKinley A Martinac B 《European biophysics journal : EBJ》2005,34(5):403-412
The periplasmic loop of MscL, the mechanosensitive channel of large conductance, acts as a spring resisting the opening of the channel. Recently, a high-throughput functional screening of a range of MscL structural mutants indicated that the substitution of residue glutamine (Q) 65 with arginine (R) or leucine (L) leads to a wild-type (WT)-like and a loss-of-function (LOF) phenotype, respectively (Maurer and Dougherty J. Biol. Chem. 278(23):21076–21082, 2003). We used electron paramagnetic resonance (EPR) spectroscopy, single-channel recording and in vivo experiments to investigate further the effect of R and L mutation of Q65 on the gating mechanism of MscL. Structural analysis of Q65R and Q65L was carried out by coupling the site-directed spin labeling (SDSL) with EPR spectroscopy. A SDSL cysteine mutant of the isoleucine 24 residue (I24C-SL) in the first transmembrane domain, TM1, of MscL served as a reporter residue in EPR experiments. This was due to its strong spin–spin interaction with the neighboring I24C-SL residues in the MscL channel pentamer (Perozo et al.Nature 418:942–948, 2002). The effects of bilayer incorporation of lysophosphatidylcholine on the MscL mutants were also investigated. Functional analysis was carried out using patch-clamp recordings from these mutants and WT MscL reconstituted into artificial liposomes. Although our data are largely in agreement with the high-throughput mutational analysis of Maurer and Dougherty, this study shows that Q65R and Q65L form functional channels and that these mutations lead to partial gain-of-function (GOF) and LOF mutation, respectively. Overall, our study confirms and advances the notion that the periplasmic loop plays a role in setting the channel mechanosensitivity.A Proceeding of the 28th Annual Meeting of the Australian Society for Biophysics 相似文献
67.
van der Kraan MI van der Made C Nazmi K van't Hof W Groenink J Veerman EC Bolscher JG Nieuw Amerongen AV 《Peptides》2005,26(11):2093-2097
LFampin 265-284, derived from bovine lactoferrin, has broad-spectrum antimicrobial activity against the yeast Candida albicans and several Gram-positive and Gram-negative bacteria. A glycine substitution scan was used to identify residues that are important for its candidacidal activity. Each single substitution of a positively charged residue led to considerable reduction in candidacidal activity, for each residue to a different extent. Substitution within the helix-facilitating N-terminal sequence DLIW had less severe effect; substitution of Ile and Trp led to a somewhat reduced potency. No substantial effects were found on the propensity to adopt a helical structure or to bind to C. albicans cells. 相似文献
68.
Olicard C Didier Y Marty C Bourgougnon N Renault T 《Diseases of aquatic organisms》2005,67(1-2):141-147
Mortalities related to the detection of Ostreid Herpesvirus 1 (OsHV-1) have been previously reported in France among larvae and spat of the Pacific oyster Crassostrea gigas. Adult oysters appear less sensitive to herpesvirus infections, although OsHV-1 has been detected in adults without signs of disease or mortality. This suggests that the virus is able to persist in its host and that adult oysters may be able to control OsHV-1 infection. Little is known about antiviral substances in invertebrates. The present work concerns the research of antiviral substances in adult oyster C. gigas, where putative antiviral activities were monitored using 3 strategies: (1) in metabolites with variable polarity, (2) in peptidic extracts and (3) in crude haemolymph. In vitro antiviral assays were based on inhibition of Herpes simplex virus type 1 (HSV-1) replication in Vero cell monolayers. All extracts presented no cytotoxicity. Antiviral activity was detected in the fresh filtered haemolymph (EC50:425 microg ml(-1)) and seasonal variation of the haemolymph antiviral activity was monitored. 相似文献
69.
Pauwels B Korst AE Andriessen V Baay MF Pattyn GG Lambrechts HA Pooter CM Lardon F Vermorken JB 《Radiation research》2005,164(5):642-650
Gemcitabine has excellent radiosensitizing properties, as shown in both preclinical and clinical studies. Radiosensitization correlated with the early S-phase block of gemcitabine. In the present study, we investigated the role of TP53 in the radiosensitizing effect of gemcitabine. Isogenic A549 cells differing in TP53 status were treated with gemcitabine during the 24 h prior to irradiation. Cell survival was determined 7 days after irradiation by the sulforhodamine B test. In addition, cell cycle perturbation was determined by flow cytometry and TP53 expression by Western blot analysis. Gemcitabine caused a concentration-dependent radiosensitizing effect in all cell lines. Transformed A549 cells were less sensitive to the cytotoxic effect of gemcitabine. The cell cycle arrest early in the S phase was dependent on the drug dose but was comparable in the different cell lines and was not related to functional TP53. Using isogenic cell lines, we have shown that neither TP53 status nor the transfection procedure influenced the radiosensitizing effect of gemcitabine. Since both the radiosensitizing effect at equitoxic concentrations and the cell cycle effect of gemcitabine were independent of TP53 expression, it is likely that TP53 protein does not play a crucial role in the radiosensitizing mechanism of gemcitabine. 相似文献
70.
Defence of Rhizobium etli bacteroids against oxidative stress involves a complexly regulated atypical 2-Cys peroxiredoxin 总被引:1,自引:0,他引:1
Dombrecht B Heusdens C Beullens S Verreth C Mulkers E Proost P Vanderleyden J Michiels J 《Molecular microbiology》2005,55(4):1207-1221
In general, oxidative stress, the consequence of an aerobic lifestyle, induces bacterial antioxidant defence enzymes. Here we report on a peroxiredoxin of Rhizobium etli, prxS, strongly expressed under microaerobic conditions and during the symbiotic interaction with Phaseolus vulgaris. The microaerobic induction of the prxS-rpoN2 operon is mediated by the alternative sigma factor RpoN and the enhancer-binding protein NifA. The RpoN-dependent promoter is also active under low-nitrogen conditions through the enhancer-binding protein NtrC. An additional symbiosis-specific weak promoter is located between prxS and rpoN2. Constitutive expression of prxS confers enhanced survival and growth to R. etli in the presence of H2O2. Single prxS mutants are not affected in their symbiotic abilities or defence response against oxidative stress under free-living conditions. In contrast, a prxS katG double mutant has a significantly reduced (>40%) nitrogen fixation capacity, suggesting a functional redundancy between PrxS and KatG, a bifunctional catalase-peroxidase. In vitro assays demonstrate the reduction of PrxS protein by DTT and thioredoxin. PrxS displays substrate specificity towards H2O2 (Km = 62 microM) over alkyl hydroperoxides (Km > 1 mM). Peroxidase activity is abolished in both the peroxidatic (C56) and resolving (C156) cysteine PrxS mutants, while the conserved C81 residue is required for proper folding of the protein. Resolving of the R. etli PrxS peroxidatic cysteine is probably an intramolecular process and intra- and intersubunit associations were observed. Taken together, our data support, for the first time, a role for an atypical 2-Cys peroxiredoxin against oxidative stress in R. etli bacteroids. 相似文献