Effects of the Mammalian Target of Rapamycin Inhibitor Everolimus on Hepatitis C Virus Replication In Vitro and In Vivo.

BACKGROUND: The influence of immunosuppressants on hepatitis C virus (HCV) re-infection after liver transplantation, particularly mammalian target of rapamycin inhibitors, remains unclear. The aim of our study was to analyze the influence of everolimus (EVR) on HCV replication activity in the context of underlying molecular mechanisms, with focus on the pro-myelocytic leukemia protein (PML). METHODS: HCV viral load was recorded in 40 patients with post-transplant HCV re-infection before and 8 weeks after introduction of EVR. An HCV cell culture replicon system for genotype (GT) 1b, GT2b, and GT3a was used to compare the influence of EVR on HCV replication for the respective genotypes in vitro. Fluorescence-activated cell-sorting analysis was used to test for effects on cell proliferation. PML expression was silenced with the use of small hairpin RNA constructs, and PML expression was quantified by means of quantitative real-time polymerase chain reaction. RESULTS: In patients with HCV, the viral load of GT1a and GT1b was hardly affected by EVR, whereas the viral load was reduced in patients with GT2a (P ≤ .0001) or GT3 infection (P ≤ .05). In vitro EVR impairs HCV replication activity of GT2a and GT3a up to 60% (P ≤ .0005), whereas in GT1b cells, HCV replication activity is increased by 50% (P ≤ .005). Replicon cell viability was not impaired. HCV replication activity is impaired in the absence of PML, which can be reversed by overexpression of PML isoforms. Furthermore, in the absence of PML, the effect of EVR on HCV replication activity is nearly abrogated. CONCLUSIONS: The mammalian target of rapamycin inhibitor EVR influences HCV replication via PML. The herein presented results suggest a genotype-dependent benefit for an EVR-based immunosuppressive regimen in patients with GT2a or GT3 re-infection after liver transplantation.

AAV2/8-humanFOXP3 gene therapy shows robust anti-atherosclerosis efficacy in LDLR-KO mice on high cholesterol diet.

Inflammation is a key etiologic component in atherogenesis. Previously we demonstrated that adeno-associated virus (AAV) 2/8 gene delivery of Netrin1 inhibited atherosclerosis in the low density lipoprotein receptor knockout mice on high-cholesterol diet (LDLR-KO/HCD). One important finding from this study was that FOXP3 was strongly up-regulated in these Netrin1-treated animals, as FOXP3 is an anti-inflammatory gene, being the master transcription factor of regulatory T cells. These results suggested that the FOXP3 gene might potentially be used, itself, as an agent to limit atherosclerosis. To test this hypothesis AAV2/8 (AAV)/hFOXP3 or AAV/Neo (control) gene therapy virus were tail vein injected into the LDLR-KO/HCD animal model. It was found that hFOXP3 gene delivery was associated with significantly lower HCD-induced atherogenesis, as measured by larger aortic lumen cross sectional area, thinner aortic wall thickness, and lower aortic systolic blood velocity compared with Neo gene-HCD-treated controls. Moreover these measurements taken from the hFOXP3/HCD-treated animals very closely matched those measurements taken from the normal diet (ND) control animals. These data strongly suggest that AAV/hFOXP3 delivery gave a robust anti-atherosclerosis therapeutic effect and further suggest that FOXP3 be examined more stringently as a therapeutic gene for clinical use.

Proton range monitoring with in-beam PET: Monte Carlo activity predictions and comparison with cyclotron data.

GOAL: Proton treatment monitoring with Positron-Emission-Tomography (PET) is based on comparing measured and Monte Carlo (MC) predicted ß(+) activity distributions. Here we present PET ß(+) activity data and MC predictions both during and after proton irradiation of homogeneous PMMA targets, where protons were extracted from a cyclotron. METHODS AND MATERIALS: PMMA phantoms were irradiated with 62 MeV protons extracted from the CATANA cyclotron. PET activity data were acquired with a 10 × 10 cm(2) planar PET system and compared with predictions from the FLUKA MC generator. We investigated which isotopes are produced and decay during irradiation, and compared them to the situation after irradiation. For various irradiation conditions we compared one-dimensional activity distributions of MC and data, focussing on Δw50%, i.e., the distance between the 50% rise and 50% fall-off position. RESULTS: The PET system is able to acquire data during and after cyclotron irradiation. For PMMA phantoms the difference between the FLUKA MC prediction and our data in Δw50% is less than 1 mm. The ratio of PET activity events during and after irradiation is about 1 in both data and FLUKA, when equal time-frames are considered. Some differences are observed in profile shape. CONCLUSION: We found a good agreement in Δw50% and in the ratio between beam-on and beam-off activity between the PET data and the FLUKA MC predictions in all irradiation conditions.

First full-beam PET acquisitions in proton therapy with a modular dual-head dedicated system.

During particle therapy irradiation, positron emitters with half-lives ranging from 2 to 20 min are generated from nuclear processes. The half-lives are such that it is possible either to detect the positron signal in the treatment room using an in-beam positron emission tomography (PET) system, right after the irradiation, or to quickly transfer the patient to a close PET/CT scanner. Since the activity distribution is spatially correlated with the dose, it is possible to use PET imaging as an indirect method to assure the quality of the dose delivery. In this work, we present a new dedicated PET system able to operate in-beam. The PET apparatus consists in two 10 cm × 10 cm detector heads. Each detector is composed of four scintillating matrices of 23 × 23 LYSO crystals. The crystal size is 1.9 mm × 1.9 mm × 16 mm. Each scintillation matrix is read out independently with a modularized acquisition system. The distance between the two opposing detector heads was set to 20 cm. The system has very low dead time per detector area and a 3 ns coincidence window, which is capable to sustain high single count rates and to keep the random counts relatively low. This allows a new full-beam monitoring modality that includes data acquisition also while the beam is on. The PET system was tested during the irradiation at the CATANA (INFN, Catania, Italy) cyclotron-based proton therapy facility. Four acquisitions with different doses and dose rates were analysed. In all cases the random to total coincidences ratio was equal or less than 25%. For each measurement we estimated the accuracy and precision of the activity range on a set of voxel lines within an irradiated PMMA phantom. Results show that the inclusion of data acquired during the irradiation, referred to as beam-on data, improves both the precision and accuracy of the range measurement with respect to data acquired only after irradiation. Beam-on data alone are enough to give precisions better than 1 mm when at least 5 Gy are delivered.

Superallowed Gamow-Teller decay of the doubly magic nucleus 100Sn.

The shell structure of atomic nuclei is associated with 'magic numbers' and originates in the nearly independent motion of neutrons and protons in a mean potential generated by all nucleons. During ß(+)-decay, a proton transforms into a neutron in a previously not fully occupied orbital, emitting a positron-neutrino pair with either parallel or antiparallel spins, in a Gamow-Teller or Fermi transition, respectively. The transition probability, or strength, of a Gamow-Teller transition depends sensitively on the underlying shell structure and is usually distributed among many states in the neighbouring nucleus. Here we report measurements of the half-life and decay energy for the decay of (100)Sn, the heaviest doubly magic nucleus with equal numbers of protons and neutrons. In the ß-decay of (100)Sn, a large fraction of the strength is observable because of the large decay energy. We determine the largest Gamow-Teller strength so far measured in allowed nuclear ß-decay, establishing the 'superallowed' nature of this Gamow-Teller transition. The large strength and the low-energy states in the daughter nucleus, (100)In, are well reproduced by modern, large-scale shell model calculations.

[Impact of preoperative pain on postoperative pain chronification. Six-month follow-up after urologic surgery]. / Einfluss präoperativer Schmerzen auf die postoperative Schmerzchronifizierung. Sechs Monate Nachbeobachtung nach urologischen Operationen.

BACKGROUND: We examined the influence of preoperative pain on postoperative pain chronification in urological patients. METHODS: Pain was determined before operation, immediately afterwards and 3 or 6 months post-surgically. Acute and chronic pain was analysed in detail with regard to severity, grade of chronification and pain intensity. We also assessed patients with the Hospital Anxiety and Depression Scale. RESULTS: Patients with preoperative pain reported postoperatively higher pain scores compared to patients without preoperative pain. Patients with higher Hospital Anxiety and Depression Scale scores reported higher pain scores for the first 7 days after the operation. Three months after surgery 51.2% of all patients and 6 months after surgery 1.2% of all patients reported about pain. CONCLUSION: Our results give evidence to the fact that preexisting pain prior to surgery has an influence on the postoperative pain course. To avoid chronification adequate therapy of the preexisting pain should be carried out.

[Postoperative pain therapy after radical prostatectomy with and without epidural analgesia]. / Postoperative Schmerztherapie nach radikaler Prostatektomie mit oder ohne epiduraler Analgesie.

Epidural analgesia for postoperative pain treatment is favored, for example, within the scope of so-called fast-track surgery, especially abdominal surgery. To improve pain care for our urological patients, we examined the quality of postoperative pain therapy with and without epidural analgesia after radical prostatectomy. After the investigation was approved by the local ethics committee, patients were questioned in detail about the pain they experienced for 7 days after radical prostatectomy. For all 7 postoperative observation days, significantly less pain was measured for patients receiving epidural analgesia compared with patients without epidural analgesia. This could be shown for the average and strongest pain intensity at rest as well as for pain during mobilization. Patients with epidural analgesia were discharged, on average, 1 day earlier. After radical prostatectomy, postoperative pain therapy with epidural analgesia seems to offer advantages with regard to the quality of analgesia and the average length of hospital stay.

[Preoperative pain in urological patients scheduled for surgical intervention : Incidence, severity and chronicity]. / Präoperative Schmerzen bei urologisch operativen Patienten : Häufigkeit, Intensität und Chronizität.

BACKGROUND: The genesis of chronic pain in urology has so far been insufficiently investigated. No investigations have focused on the occurrence of preoperative pain. We developed an epidemiological questionnaire to analyze preoperative pain. METHODS: In this questionnaire, preoperative pain in all patients scheduled for urologic surgery (n=165) was analyzed. Acute and chronic pain was analyzed as main or adjoint pain, with the registration of severity, chronification states, and duration. We registered depression and anxiety, well-being, and somatic and psychological efficiency. RESULTS: Eighty percent of the patients reported pain within the previous 12 months. Acute preoperative pain was reported by 17% of the patients and chronic pain by 64%. Significant differences in quality of life were detected between patients with or without preoperative pain. Well-being was also significantly affected in patients having pain. CONCLUSION: The pain severity and states of chronification not only explain a reduction in somatic and psychological well-being but also emphasize that preoperative pain should be identified thoroughly prior to surgery.

Body dissatisfaction and restrained eating in male juvenile and adult athletes.

Athletes of light-weight sport classes are under a constant strain to control eating and body shape, which can make them prone to develop eating disorders. In the present study, cognitive control of eating (restrained eating) and body dissatisfaction were investigated in male elite athletes of light-weight and heavy-weight classes at different ages. Body dissatisfaction was assessed under hunger and satiety. Adult light-weight rowers had extremely high scores of restrained eating and a more pronounced body dissatisfaction under hunger compared to satiety in contrast to heavy-weight rowers. Juvenile light-weight rowers had a pronounced cognitive control of eating behavior while body dissatisfaction was not affected by weight-class or hunger. The results suggest that extensive participation in a light-weight sport increases the cognitive control of eating behavior but not the disinhibition of cognitive control of eating. High levels of cognitive control of eating in the adult lightweight rowers are accompanied with body dissatisfaction under hunger but not under satiety.

Geobacter bremensis sp. nov. and Geobacter pelophilus sp. nov., two dissimilatory ferric-iron-reducing bacteria.

Two strictly anaerobic, dissimilatory ferric-iron-reducing bacteria, strains Dfr1T and Dfr2T, were isolated from freshwater mud samples with ferrihydrite as electron acceptor. Both strains also grew by reducing Mn(IV), S0 and fumarate. Electron donors used by strains Dfr1T and Dfr2T for growth with ferric iron as electron acceptor included hydrogen, formate, acetate, pyruvate, succinate, fumarate and ethanol. An affiliation with the family Geobacteraceae was revealed by comparative analysis of 165 rRNA gene sequences. Strains Dfr1T and Dfr2T shared 92.5% sequence identity and their closest known relative was Geobacter sulfurreducens, with approximately 93% sequence identity. Cultures and colonies of strains Dfr1T and Dfr2T were intensely red in colour, due to the presence of c-type cytochromes. On the basis of physiological and phylogenetic data, strain Dfr1T (= DSM 12179T = OCM 796T) is described as Geobacter bremensis sp. nov. and strain Dfr2T (= DSM 12255T = OCM 797T) as Geobacter pelophilus sp. nov.

Prosodic structure and wh-questions.

This study examines the influence of wh-gaps on the prosodic contour of spoken utterances. A previous study (Nagel, Shapiro, & Nawy, 1994) claimed that the phonological representation of a sentence containing a filler-gap dependency explicitly encodes the location of the syntactic gap. In support of this hypothesis, Nagel et al. presented evidence that the word immediately preceding a gap is lengthened and that there is a reliable increase in pitch excursion across the gap location. Our study challenges Nagel et al.'s claim. We argue that their materials confounded the presence/ absence of a gap with other factors that are known to affect intonational phrasing independently. We show that, when these factors are separated, the evidence that syntactic gaps are explicitly encoded in the phonological representation of a sentence disappears.

Rhodovulum iodosum sp. nov. and Rhodovulum robiginosum sp. nov., two new marine phototrophic ferrous-iron-oxidizing purple bacteria.

Two new strains of marine purple bacteria, N1T and N2T, were isolated from coastal sediment of the North Sea (Germany) with ferrous iron as the only electron donor for anoxygenic photosynthesis. The isolates are the first salt-dependent, ferrous-iron-oxidizing purple bacteria characterized so far. Analysis of 16S rRNA gene sequences revealed an affiliation with the genus Rhodovulum, which until now comprises only marine species. The sequence similarity of both strains was 95.2%, and their closest relative was Rhodovulum adriaticum. Like all known Rhodovulum species, the new strains had ovoid to rod-shaped cells, contained bacteriochlorophyll a and carotenoids of the spheroidene series, and were able to oxidize sulfide and thiosulfate. Like Rhodovulum adriaticum, both strains were unable to assimilate sulfate; for growth they needed a reduced sulfur source, e.g. thiosulfate. In contrast to the new strains, none of the known Rhodovulum species tested was able to oxidize ferrous iron or iron sulfide. In growth experiments, strains N1T and N2T oxidized 65 and 95%, respectively, of the ferrous iron supplied. Electron diffraction analysis revealed ferrihydrite as the main product of ferrous iron oxidation. In addition, traces of magnetite were formed. Strains N1T (= DSM 12328T) and N2T (= DSM 12329T) are described as Rhodovulum iodosum sp. nov. and Rhodovulum robiginosum sp. nov., respectively.

Enumeration and detection of anaerobic ferrous iron-oxidizing, nitrate-reducing bacteria from diverse European sediments.

Anaerobic, nitrate-dependent microbial oxidation of ferrous iron was recently recognized as a new type of metabolism. In order to study the occurrence of three novel groups of ferrous iron-oxidizing, nitrate-reducing bacteria (represented by strains BrG1, BrG2, and BrG3), 16S rRNA-targeted oligonucleotide probes were developed. In pure-culture experiments, these probes were shown to be suitable for fluorescent in situ hybridization, as well as for hybridization analysis of denaturing gradient gel electrophoresis (DGGE) patterns. However, neither enumeration by in situ hybridization nor detection by the DGGE-hybridization approach was feasible with sediment samples. Therefore, the DGGE-hybridization approach was combined with microbiological methods. Freshwater sediment samples from different European locations were used for enrichment cultures and most-probable-number (MPN) determinations. Bacteria with the ability to oxidize ferrous iron under nitrate-reducing conditions were detected in all of the sediment samples investigated. At least one of the previously described types of bacteria was detected in each enrichment culture. MPN studies showed that sediments contained from 1 x 10(5) to 5 x 10(8) ferrous iron-oxidizing, nitrate-reducing bacteria per g (dry weight) of sediment, which accounted for at most 0.8% of the nitrate-reducing bacteria growing with acetate. Type BrG1, BrG2, and BrG3 bacteria accounted for an even smaller fraction (0.2% or less) of the ferrous iron-oxidizing, nitrate-reducing community. The DGGE patterns of MPN cultures suggested that more organisms than those isolated thus far are able to oxidize ferrous iron with nitrate. A comparison showed that among the anoxygenic phototrophic bacteria, organisms that have the ability to oxidize ferrous iron also account for only a minor fraction of the population.

Technology and the business office.

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The use of biologically produced ferrihydrite for the isolation of novel iron-reducing bacteria.

Ferric iron was produced anaerobically from ferrous iron through the metabolic activity of recently described ferrous iron-oxidizing, nitrate-reducing bacteria. It was identified as poorly crystallized 2-line ferrihydrite with a particle size of 1-2 nm. This biologically produced ferrihydrite was shown to be a suitable electron acceptor for dissimilatory ferric iron-reducing bacteria in freshwater enrichment cultures, and was completely reduced to the ferrous state; no magnetite formation occurred. Geobacter metallireducens was also able to completely reduce the biologically produced ferrihydrite. These results indicate the possibility of an anaerobic, microbial cycling of iron. Using the biologically produced ferric iron, two isolates of obligately anaerobic, dissimilatory ferric iron-reducing bacteria, strains Dfr1 and Dfr2, were obtained from freshwater enrichment cultures. Analysis of 16S rRNA gene sequences revealed an affiliation with the Geobacter cluster within the family Geobacteraceae. The sequence similarity between strains Dfr1 and Dfr2 is 92.5%. The closest relative of strain Dfr1 is Geobacter sulfurreducens with 92.9%, and of strain Dfr2 Geobacter chapelleii with 93.7% sequence similarity. In addition, strains Dfr1 and Dfr2 are both able to grow by dissimilatory reduction of Mn(IV), S degree, and fumarate. Furthermore, strain Dfr2 is able to reduce akaganeite (beta-FeOOH), a more crystallized type of ferric iron oxide.

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Linking telephone and computer technologies to deliver information can help a health care organization accomplish strategic business goals: better customer service and bottom line.

Financial systems: the next generation.

Changes in health care organizations and business styles are creating demand for new financial applications that span enterprises, deliver managed care muscle to providers, mesh with CPRs and solve Year 2000 concerns.

Laser energy reaching the posterior pole during transscleral cyclophotocoagulation.

OBJECTIVE: To measure scattered laser energy reaching the posterior pole during transscleral cyclophotocoagulation. METHODS: Transscleral cyclophotocoagulation was performed on 4 cadaver eyes with Nd:YAG noncontact, Nd:YAG contact, and diode contact lasers. Energy was measured with a photodiode through a 7-mm trephined hole in the posterior pole. Average percentage power, average power, and average energy transmission were calculated. American Conference of Governmental Industrial Hygienists (ACGIH) guidelines were used to calculate allowable energy exposures for each laser. RESULTS: All 3 lasers transmitted 3% to 5% of the power to the posterior pole. The average energy transmission was 240 to 260 mJ for all lasers. The contact lasers had an average power transmission of 120 mW. The noncontact Nd:YAG laser, with shorter pulse duration, had an average power transmission of 13,000 mW, significantly greater than that of the other lasers. The ACGIH guidelines for allowable energy exposures were 93 mJ for the noncontact Nd:YAG laser, 1300 mJ for the contact Nd:YAG laser, and 440 mJ for the contact diode laser. CONCLUSIONS: Three percent to 5% of laser power delivered during cyclophotocoagulation reaches the posterior pole. Exposure energies may approach or exceed ACGIH guidelines. The clinical significance of these findings remains to be shown.