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Background: Continuous administration of levodopa-carbidopa intestinal gel (carbidopa-levodopa enteral suspension) through a percutaneous endoscopic gastrojejunostomy is a treatment option for advanced Parkinson disease (PD) patients with motor fluctuations resistant to standard oral medications. Safety data from 4 prospective studies were integrated to assess the safety of this therapy.Methods: Safety data from 4 studies were summarized using 2 overlapping data sets, permitting the separation of procedure/device-associated (n = 395) from non-procedure/device adverse events (n = 412).Results: At the data cutoff, median exposure to levodopa-carbidopa intestinal gel was 911 days (range, 1-1980 days) with 963 total patient-years of exposure. Procedure/device adverse events occurred in 300 patients (76%), and serious adverse events occurred in 68 (17%); most frequently reported procedure/device adverse events and serious adverse events were complications of device insertion (41% and 8%, respectively) and abdominal pain (36% and 4%, respectively). Non-procedure/device adverse events occurred in 92% (379), with most frequently reported being insomnia (23%) and falls (23%); 42% (171) had non-procedure/device serious adverse events, with most frequently reported being pneumonia (5%) and PD symptoms (2%). Adverse events led to discontinuation in 17% (72), most frequently because of complication of device insertion (2.4%). There were 34 treatment-emergent deaths (8.3%) in the overlapping data sets, 2 of which (0.5%) were considered "possibly related" to the treatment system.Conclusion: In the largest collection of levodopa-carbidopa intestinal gel safety data from prospective clinical studies, procedure/device events were frequently reported and occasionally life threatening. Most non-procedure/device events were typical for levodopa treatment and an elderly population. These factors combined with high treatment efficacy led to a relatively low discontinuation rate in advanced PD patients. [ABSTRACT FROM AUTHOR]

Hepatitis E virus (HEV) infection is a cause of hepatitis in humans worldwide and has been associated with a case-fatality rate of up to 30% in pregnant women. Recently, persistent and chronic HEV infections have been recognized as a serious clinical problem, especially in immunocompromised individuals. To date, there are no FDA-approved HEV-specific antiviral drugs. In this study, we evaluated antisense peptide-conjugated morpholino oligomers (PPMO) designed against HEV genomic sequences as potential HEV-specific antiviral compounds. Two genetically-distinct strains of human HEV, genotype 1 Sar55 and genotype 3 Kernow-C1, isolated from patients with acute and chronic hepatitis, respectively, were used to evaluate inhibition of viral replication by PPMO in liver cells. The anti-HEV PPMO produced a significant reduction in the levels of HEV RNA and capsid protein, indicating effective inhibition of HEV replication. PPMO HP1, which targets a highly conserved sequence in the start site region of ORF1, was also effective against the genotype 3 Kernow-C1 strain in stably-infected HepG2/C3A liver cells. The antiviral activity observed was specific, dose-responsive and potent, suggesting that further exploration of PPMO HP1 as a potential HEV-specific antiviral agent is warranted. [ABSTRACT FROM AUTHOR]

Summary: Type I interferons (IFN-I) are essential antiviral cytokines produced upon microbial infection. IFN-I elicits this activity through the upregulation of hundreds of IFN-I-stimulated genes (ISGs). The full breadth of ISG induction demands activation of a number of cellular factors including the IκB kinase epsilon (IKKε). However, the mechanism of IKKε activation upon IFN receptor signaling has remained elusive. Here we show that TRIM6, a member of the E3-ubiquitin ligase tripartite motif (TRIM) family of proteins, interacted with IKKε and promoted induction of IKKε-dependent ISGs. TRIM6 and the E2-ubiquitin conjugase UbE2K cooperated in the synthesis of unanchored K48-linked polyubiquitin chains, which activated IKKε for subsequent STAT1 phosphorylation. Our work attributes a previously unrecognized activating role of K48-linked unanchored polyubiquitin chains in kinase activation and identifies the UbE2K-TRIM6-ubiquitin axis as critical for IFN signaling and antiviral response. [ABSTRACT FROM AUTHOR]

Ferroelectrics have recently attracted attention as a candidate class of materials for use in photovoltaic devices, and for the coupling of light absorption with other functional properties. In these materials, the strong inversion symmetry breaking that is due to spontaneous electric polarization promotes the desirable separation of photo-excited carriers and allows voltages higher than the bandgap, which may enable efficiencies beyond the maximum possible in a conventional p-n junction solar cell. Ferroelectric oxides are also stable in a wide range of mechanical, chemical and thermal conditions and can be fabricated using low-cost methods such as sol-gel thin-film deposition and sputtering. Recent work has shown how a decrease in ferroelectric layer thickness and judicious engineering of domain structures and ferroelectric-electrode interfaces can greatly increase the current harvested from ferroelectric absorber materials, increasing the power conversion efficiency from about 10−4 to about 0.5 per cent. Further improvements in photovoltaic efficiency have been inhibited by the wide bandgaps (2.7-4 electronvolts) of ferroelectric oxides, which allow the use of only 8-20 per cent of the solar spectrum. Here we describe a family of single-phase solid oxide solutions made from low-cost and non-toxic elements using conventional solid-state methods: [KNbO3]1 − x[BaNi1/2Nb1/2O3 − δ]x (KBNNO). These oxides exhibit both ferroelectricity and a wide variation of direct bandgaps in the range 1.1-3.8 electronvolts. In particular, the x = 0.1 composition is polar at room temperature, has a direct bandgap of 1.39 electronvolts and has a photocurrent density approximately 50 times larger than that of the classic ferroelectric (Pb,La)(Zr,Ti)O3 material. The ability of KBNNO to absorb three to six times more solar energy than the current ferroelectric materials suggests a route to viable ferroelectric semiconductor-based cells for solar energy conversion and other applications. [ABSTRACT FROM AUTHOR]

Dengue virus has emerged as a global health threat to over one-third of humankind. As a positive-strand RNA virus, dengue virus relies on the host cell metabolism for its translation, replication, and egress. Therefore, a better understanding of the host cell metabolic pathways required for dengue virus infection offers the opportunity to develop new approaches for therapeutic intervention. In a recently described screen of known drugs and bioactive molecules, we observed that methotrexate and floxuridine inhibited dengue virus infections at low micromolar concentrations. Here, we demonstrate that all serotypes of dengue virus, as well as West Nile virus, are highly sensitive to both methotrexate and floxuridine, whereas other RNA viruses (Sindbis virus and vesicular stomatitis virus) are not. Interestingly, flavivirus replication was restored by folinic acid, a thymidine precursor, in the presence of methotrexate and by thymidine in the presence of floxuridine, suggesting an unexpected role for thymidine in flavivirus replication. Since thymidine is not incorporated into RNA genomes, it is likely that increased thymidine production is indirectly involved in flavivirus replication. A possible mechanism is suggested by the finding that p53 inhibition restored dengue virus replication in the presence of floxuridine, consistent with thymidine-less stress triggering p53-mediated antiflavivirus effects in infected cells. Our data reveal thymidine synthesis pathways as new and unexpected therapeutic targets for antiflaviviral drug development. [ABSTRACT FROM AUTHOR]

Summary: Drosophila embryo dorsoventral (DV) polarity is defined by serine protease activity in the perivitelline space (PVS) between the embryonic membrane and the inner layer of the eggshell []. Gastrulation Defective (GD) cleaves and activates Snake (Snk). Activated Snk cleaves and activates Easter (Ea), exclusively on the ventral side of the embryo []. Activated Ea then processes Spätzle (Spz) into the activating ligand for Toll, a transmembrane receptor that is distributed throughout the embryonic plasma membrane []. Ventral activation of Toll depends upon the activity of the Pipe sulfotransferase in the ventral region of the follicular epithelium that surrounds the developing oocyte []. Pipe transfers sulfate residues to several protein components of the inner vitelline membrane layer of the eggshell []. Here we show that GD protein becomes localized in the ventral PVS in a Pipe-dependent process. Moreover, ventrally concentrated GD acts to promote the cleavage of Ea by Snk through an extracatalytic mechanism that is distinct from GD''s proteolytic activation of Snk. Together, these observations illuminate the mechanism through which spatially restricted sulfotransferase activity in the developing egg chamber leads to localization of serine protease activity and ultimately to spatially specific activation of the Toll receptor in the Drosophila embryo. [Copyright &y& Elsevier]

Myeloid Sarcoma is a rare tumor composed of myeloblasts occurring at an extramedullary site like bones, or various soft tissues. Myeloid sarcoma may involve the gastrointestinal tract very rarely either solitarily, or occurring simultaneously with acute myeloid leukemia. Its diagnosis is challenging and needs biopsy and immunohistochemical staining. We are describing a case of myeloid sarcoma which presented as a painful anal ulcer mimicking an atypical fissure. Its appearance resembled crohn's disease on sigmoidoscopy. A biopsy of the ulcer along with histochemical staining led to the diagnosis of myeloid sarcoma. Our case demonstrates the need for aggressive evaluation of any common gastrointestinal complaint with an atypical presentation. [ABSTRACT FROM AUTHOR]

The establishment of dorsal-ventral (DV) polarity in the Drosophila embryo depends upon a localized signal that is generated in the perivitelline space of the egg through the action of a serine proteolytic cascade. Spatial regulation of this pathway is determined by the expression of the pipe gene in a subpopulation of ventral follicle cells in the developing egg chamber. The Pipe protein exhibits homology to vertebrate glycosaminoglycan sulfotransferases. In a previous study, we demonstrated that embryonic DV polarity depends upon the sulfotransferase activity of Pipe. Surprisingly, however, our results also indicated that formation of the embryonic DV axis does not require the synthesis of the high-energy sulfate donor, 3′-phosphoadenosine 5′-phosphosulfate (PAPS) in the follicle cells in which Pipe is presumed to function. Here, we resolve this apparent paradox by demonstrating that dorsalized embryos are only produced by egg chambers in which both germline and follicle cells lack PAPS synthetase activity. Thus, PAPS produced either in the germline or in the follicular epithelium can support the requirement for Pipe sulfotransferase activity in embryonic DV patterning. This finding indicates the existence of a conduit for the movement of PAPS between the germline and the follicle cells, which highlights a previously unappreciated mechanism of soma/germline cooperation affecting pattern formation. [ABSTRACT FROM AUTHOR]

Purpose: To determine whether different doses and dosing regimens of intravitreal vascular endothelial growth factor (VEGF) Trap-Eye are superior to focal/grid photocoagulation in eyes with diabetic macular edema (DME). Design: Multicenter, randomized, double-masked, phase 2 clinical trial. Participants: A total of 221 diabetic patients with clinically significant macular edema involving the central macula. Methods: Patients were assigned to 1 of 5 treatment regimens: 0.5 mg VEGF Trap-Eye every 4 weeks; 2 mg VEGF Trap-Eye every 4 weeks; 2 mg VEGF Trap-Eye for 3 initial monthly doses and then every 8 weeks; 2 mg VEGF Trap-Eye for 3 initial monthly doses and then on an as-needed (PRN) basis; or macular laser photocoagulation. Assessments were completed at baseline and every 4 weeks thereafter. Main Outcome Measures: Mean change in visual acuity and central retinal thickness (CRT) at 24 weeks. Results: Patients in the 4 VEGF Trap-Eye groups experienced mean visual acuity benefits ranging from +8.5 to +11.4 Early Treatment of Diabetic Retinopathy Study (ETDRS) letters versus only +2.5 letters in the laser group (P ≤ 0.0085 for each VEGF Trap-Eye group vs. laser). Gains from baseline of 0+, 10+, and 15+ letters were seen in up to 93%, 64%, and 34% of VEGF Trap-Eye groups versus up to 68%, 32%, and 21% in the laser group, respectively. Mean reductions in CRT in the 4 VEGF Trap-Eye groups ranged from −127.3 to −194.5 μm compared with only −67.9 μm in the laser group (P = 0.0066 for each VEGF Trap-Eye group vs. laser). VEGF Trap-Eye was generally well tolerated. Ocular adverse events in patients treated with VEGF Trap-Eye were generally consistent with those seen with other intravitreal anti-VEGF agents. Conclusions: Intravitreal VEGF Trap-Eye produced a statistically significant and clinically relevant improvement in visual acuity when compared with macular laser photocoagulation in patients with DME. Financial Disclosure(s): Proprietary or commercial disclosure may be found after the references. [Copyright &y& Elsevier]

Abstract: Porcine reproductive and respiratory syndrome (PRRS) causes substantial economic losses to the swine industry in many countries, and current control strategies are inadequate. Previously, we explored the strategy of using peptide-conjugated phosphorodiamidate morpholino oligomers (PPMOs) to inhibit PRRS virus (PRRSV) replication. PPMOs are nuclease-resistant and single-stranded DNA analogs containing a modified backbone conjugated to a cell-penetrating peptide and can act as antisense agents through steric blockade of complementary messenger RNA. A PPMO (designated 5UP2) targeting highly conserved sequence in the 5′-terminal region of the PRRSV genome was found to produce multi-log10 inhibition of PRRSV replication in cultured cells. To evaluate 5UP2 in vivo, we here administrated the PPMO to 3-week-old piglets via intranasal instillation at 24h before, and 2 and 24h after infection with PRRSV (strain VR2385). Blood samples were collected at 6, 10 and 14days post-infection (dpi) for detection of PRRSV RNA and antibodies. Necropsy was performed at 14dpi. Monitoring weight gain in all piglet groups throughout the experiment indicated that PPMO was well tolerated at the doses used. PPMO 5UP2 treatment significantly reduced PRRSV viremia at 6dpi. On day 14, piglets receiving 5UP2 had significantly less interstitial pneumonia and lower level of anti-PRRSV antibodies than untreated piglets. In alveolar macrophages isolated at the time of necropsy, the expression of antiviral genes in PPMO-treated piglets was elevated in comparison with untreated. This study provides further data indicating that the 5UP2 PPMO can be considered a candidate component for a novel PRRS control strategy. [Copyright &y& Elsevier]

Abstract: Background context: Elective lumbar discectomy among injured workers is a prevalent spine surgery that often requires a lengthy rehabilitation. It is important to determine presurgical biopsychosocial predictors of compensation and medical costs in such patients. Purpose: To determine if presurgical biopsychosocial variables are predictive of compensation and medical costs in a cohort of Utah patients who have undergone open or microlumbar discectomy that are receiving workers'' compensation. Study design/setting: A retrospective cohort study consisting of a review of presurgical medical records and accrued medical and compensation costs. Patient sample: A consecutive sample of 266 compensated workers from Utah who had undergone either open discectomy or microlumbar discectomy from 1994 to 2000. All patients were at least 2 years postsurgery at the time of follow-up. Outcome measures: Total accrued medical, compensation, and aggregate costs. Methods: A retrospective review of presurgical biopsychosocial variables and total accrued medical, compensation, and aggregate costs. Results: Presurgical variables were statistically significantly correlated with medical and compensation costs. Multiple linear regression models accounted for 31% of variation in compensation costs, 32% in medical costs, and 43% in total aggregate costs. Conclusions: Presurgical biopsychosocial variables are important predictors of compensated lumbar discectomy costs. Medical cost control programs might benefit from identifying biopsychosocial variables related to increased costs. [Copyright &y& Elsevier]

Objectives. To examine the association of engagement in cognitively stimulating activities with cognitive and functional decline in a population-based sample of incident Alzheimer's disease (AD). Method. After diagnosis, 187 participants (65% females) were followed semiannually for a mean 2.7 (SD = 0.4) years. Mean age and education were 84.6 (SD = 5.8) and 13.2 (SD = 2.9) years. Caregivers enumerated cognitively stimulating leisure activities via the Lifestyle Activities Questionnaire. Cognition was assessed using the Mini-Mental State Examination and functional ability via the Clinical Dementia Rating sum of boxes. Linear mixed models tested the association between stimulating activities and change over time in each outcome. Covariates were demographic factors, estimated premorbid IQ, presence/absence of the APOE ε4 allele, duration of dementia, level of physical activity, and general health. Results. At initial assessment, 87% of participants were engaged in one or more stimulating activities, with mean (SD) activities = 4.0 (3.0). This number declined to 2.4 (2.0) at the final visit. There was a statistical interaction between dementia duration and number of activities in predicting rate of cognitive decline (p = .02) and overall functional ability (p = .006). Discussion. Active involvement in cognitively stimulating pursuits may be beneficial for persons with AD. [ABSTRACT FROM PUBLISHER]

Cardiomyocyte apoptosis is a hallmark of coxsackievirus B3 (CVB3)-induced myocarditis. We used cardiomyocytes and HeLa cells to explore the cellular response to CVB3 infection, with a focus on pathways leading to apoptosis. CVB3 infection triggered endoplasmic reticulum (ER) stress and differentially regulated the three arms of the unfolded protein response (UPR) initiated by the proximal ER stress sensors ATF6a (activating transcription factor 6a), IRE1-XBP1 (X box binding protein 1), and PERK (PKR-like ER protein kinase). Upon CVB3 infection, glucose-regulated protein 78 expression was upregulated, and in turn ATF6a and XBP1 were activated via protein cleavage and mRNA splicing, respectively. UPR activity was further confirmed by the enhanced expression of UPR target genes ERdj4 and EDEM1. Surprisingly, another UPRassociated gene, p58IPK, which often is upregulated during infections with other types of viruses, was downregulated at both mRNA and protein levels after CVB3 infection. These findings were observed similarly for uninfected Tet-On HeLa cells induced to overexpress ATF6a or XBP1. In exploring potential connections between the three UPR pathways, we found that the ATF6a-induced downregulation of p58IPK was associated with the activation of PKR (PERK) and the phosphorylation of eIF2α, suggesting that p58IPK, a negative regulator of PERK and PKR, mediates cross-talk between the ATF6a/IRE1-XBP1 and PERK arms. Finally, we found that CVB3 infection eventually produced the induction of the proapoptoic transcription factor CHOP and the activation of SREBP1 and caspase-12. Taken together, these data suggest that CVB3 infection activates UPR pathways and induces ER stress-mediated apoptosis through the suppression of P58IPK and induction/ activation of CHOP, SREBP1, and caspase-12. [ABSTRACT FROM AUTHOR]

Historical reviews suggest that tanning first became fashionable in the 1920s or 1930s. To quantitatively and qualitatively examine changes in tanning attitudes portrayed in the popular women's press during the early 20th century, we reviewed summer issues of Vogue and Harper's Bazaar for the years 1920, 1927, 1928, and 1929. We examined these issues for articles and advertisements promoting skin tanning or skin bleaching and protection. We found that articles and advertisements promoting the fashionable aspects of tanned skin were more numerous in 1928 and 1929 than in 1927 and 1920, whereas those promoting pale skin (by bleaching or protection) were less numerous. These findings demonstrate a clear shift in attitudes toward tanned skin during this period. [ABSTRACT FROM AUTHOR]

A primary objective of the ICEFISH 2004 cruise was to collect and study notothenioid fishes from remote localities in the Atlantic sector of the Southern Ocean. Nearly 1 month was devoted to bottom trawling for fishes on the shelf and upper slope (to 1,000 m) areas around Shag Rocks, South Georgia, South Sandwich Islands and Bouvetøya. The focus was on the latter two locations, because their faunas are more poorly known. Eight species were collected at Shag Rocks with Patagonotothen guntheri most abundant; 17 at South Georgia with Lepidonotothen nudifrons, L. larseni and Gobionotothen gibberifrons most abundant; 13 at the South Sandwich Islands with L. larseni, L. nudifrons and G. gibberifrons most abundant; and 11 at Bouvetøya with L. larseni, Macrourus holotrachys and L. squamifrons most abundant. Ten new locality records were established: Shag Rocks (1), South Georgia (1), South Sandwich Islands (5), South Sandwich Trench at 5,350 m (1) and Bouvetøya (2). Total known demersal fish diversity on the shelf and upper slope at Shag Rocks/South Georgia, South Sandwich Islands and Bouvetøya is 42, 31 and 17 species, respectively. To examine population structure in the four most abundant notothenioids at Bouvetøya ( L. larseni, L. squamifrons, Notothenia coriiceps and Chaenocephalus aceratus), we examined the ND2 portion of mitochondrial DNA. Chaenocephalus aceratus, N. coriiceps and L. larseni exhibited no significant genetic differentiation in comparison with samples from localities in the Scotia Sea and the Antarctic Peninsula. However, L. squamifrons showed significant genetic differentiation between the South Shetlands and Bouvetøya populations ( F ST = 0.189, P = 0.015). Thus, these data combined with previous studies of two other notothenioids suggest that five of the six notothenioid species at Bouvetøya are not genetically differentiated from other localities in the Atlantic sector of the Southern Ocean. The location of Bouvetøya within the Antarctic Circumpolar Current and the long (1–2 years) pelagic stages of the notothenioids at Bouvetøya may be at least partly responsible for this genetic homogeneity. [ABSTRACT FROM AUTHOR]

Abstract: Charge neutral steric block oligonucleotide analogues, such as peptide nucleic acids (PNA) or phosphorodiamidate morpholino oligomers (PMO), have promising biological and pharmacological properties for antisense applications, such as for example in mRNA splicing redirection. However, cellular uptake of free oligomers is poor and the utility of conjugates of PNA or PMO to cell penetrating peptides (CPP), such as Tat or Penetratin, is limited by endosomal sequestration. Two new families of arginine-rich CPPs named (R-Ahx-R)4 AhxB and R6Pen allow efficient nuclear delivery of splice correcting PNA and PMO at micromolar concentrations in the absence of endosomolytic agents. The in vivo efficacy of (R-Ahx-R)4 AhxB PMO conjugates has been demonstrated in mouse models of Duchenne muscular dystrophy and in various viral infections. [Copyright &y& Elsevier]

BACKGROUND Operative notes can be generated electronically by manual input of the entire note, free-form oral dictation, or using either an electronic template or a template for dictation. There are few studies that have directly compared these modalities in terms of speed, accuracy, and completeness. OBJECTIVE The objective was to determine whether electronic templates are more efficient and reduce errors compared to free-form oral dictation for the completion of Mohs micrographic surgery operative notes. METHODS Operative notes for 110 consecutive Mohs micrographic surgery cases were completed either by oral dictation or by electronic template. The time to dictate or complete the template was recorded for each note. Notes were subsequently edited, recording the number and type of errors as well as the time required to edit each note. RESULTS Compared with dictation, operative notes completed with the electronic template had fewer errors (5.8% vs. 81%), took less time to complete (175.5 seconds vs. 240.0 seconds), took less time to review and edit (41.6 seconds vs. 201.1 seconds), and were completed and signed in a more timely fashion (0.115 days vs. 20.7 days). CONCLUSION Electronic templates are a more accurate and rapid method compared to free-form oral dictation for the completion of Mohs micrographic surgery operative notes and have the advantage of being immediately available to review and sign. [ABSTRACT FROM AUTHOR]

The filoviruses Marburg virus and Ebola virus (EBOV) quickly outpace host immune responses and cause hemorrhagic fever, resulting in case fatality rates as high as 90% in humans and nearly 100% in nonhuman primates. The development of an effective therapeutic for EBOV is a daunting public health challenge and is hampered by a paucity of knowledge regarding filovirus pathogenesis. This report describes a successful strategy for interfering with EBOV infection using antisense phosphorodiamidate morpholino oligomers (PMOs). A combination of EBOV-specific PMOs targeting sequences of viral mRNAs for the viral proteins (VPs) VP24, VP35, and RNA polymerase L protected rodents in both pre- and post-exposure therapeutic regimens. In a prophylactic proof-of-principal trial, the PMOs also protected 75% of rhesus macaques from lethal EBOV infection. The work described here may contribute to development of designer, "druggable" countermeasures for filoviruses and other microbial pathogens. [ABSTRACT FROM AUTHOR]

The article discusses the study "Misspecified Cells Die by an Active Gene-Directed Process, and Inhibition of This Death Results in Cell Fate Transformation in Drosophila," by Christian Werz, Tom V. Lee et al. Incorrectly specified cells often result to cell death or are transformed to adopt a different cell fate during development. Large numbers of mis-specified cells die synchronously in many developmental mutants in Drosophila. Upregulation of hid occurs in oskar mutants and other segmentation mutants.

Background. In human immunodeficiency virus (HIV)-infected patients, fluconazole prophylaxis is associated with reductions in the rate of fungal infection. However, concerns exist with regard to the use of fluconazole prophylaxis and the risk of development of fluconazole treatment-refractory infections. Methods. We performed a randomized, open-label trial that compared oral fluconazole given continuously (200 mg 3 times weekly', the "continuous fluconazole arm") with fluconazole that was provided only for episodes of orophayngeal candidiasis (OPC) or esophageal candidiasis (EC) (the "episodic fluconazole arm") in HIV-infected persons with CD4+ T cell counts of <150 cells/mm³ and a history of OPC. The primary study end point was the time to development of fluconazole-refractory OPC or EC, which was defined as lack of response to 200 mg fluconazole given daily for 14 or 21 days, respectively. Results. A total of 413 subjects were randomized to receive continuous fluconazole, and 416 were randomized to receive episodic fluconazole. After 42 months, 17 subjects (4.1%) in the continuous fluconazole arm developed fluconazole-refractory OPC or EC infections, compared with 18 subjects (4.3%) in the episodic fluconazole arm, with no difference between treatment arms with regard to the time to development of a fluconazole-refractory infection within 24 months (P = .88, by log-rank test) or before the end of the study (P = .97, by the log-rank test). Continuous fluconazole therapy was associated with fewer cases of OPC or EC (0.29 vs. 1.08 episodes per patient-year; P <.0001) and fewer invasive fungal infections (15 vs. 28 episodes; P = .04, by χ² test), but not with improved survival, compared with episodic fluconazole therapy. Conclusion. Continuous fluconazole therapy is not associated with significant risk of fluconazole-refractory OPC or EC, compared with episodic fluconazole therapy, in HIV-infected patients with access to active antiretroviral therapy. [ABSTRACT FROM AUTHOR]

The Drosophila pipe gene encodes ten related proteins that exhibit amino acid sequence similarity to vertebrate heparan sulfate 2-O-sulfotransferase. One of the Pipe isoforms, which is expressed in the ventral follicular epithelium, is a key determinant of embryonic dorsoventral polarity, suggesting that Pipe-mediated sulfation of a heparan sulfate proteoglycan provides a spatial cue for dorsoventral axis formation. We used several approaches to investigate this possibility in the work described here. We determined the nucleotide alterations in 11 different pipe alleles. Ten of the mutations specifically affect the pipe isoform that is expressed in the ovary. Among these ten mutations, two alter an amino acid in the putative binding site for 3′-phosphoadenosine 5′-phosphosulfate, the universal sulfate donor. Using Alcian Blue, a histochemical stain that detects sulfated glycans, we observed a novel, pipe-dependent macromolecule in the embryonic salivary glands. Genes known to participate in the formation of heparan sulfate in Drosophila are not required for the production of this material. To investigate whether a heparan sulfate proteoglycan is involved in pipe function in dorsoventral patterning, we generated females carrying follicle cell clones mutant for heparan sulfate synthesis-related genes. Embryos from follicles with mutant clones did not exhibit a dorsalized phenotype. Taken together, our data provide evidence that Pipe acts as a sulfotransferase, but argue against the hypothesis that the target of Pipe is a heparan sulfate glycosaminoglycan. [ABSTRACT FROM AUTHOR]

Five dengue (DEN) virus-specific R5F2R4 peptide-conjugated phosphorodiamidate morpholino oligomers (P4-PMOs) were evaluated for their ability to inhibit replication of DEN virus serotype 2 (DEN-2 virus) in mammalian cell culture. Initial growth curves of DEN-2 virus 16681 were obtained in Vero cells incubated with 20 µM P4-PMO compounds. At 6 days after infection, a P4-PMO targeting the 3′-terminal nucleotides of the DEN-2 virus genome and a random-sequence P4-PMO showed relatively little suppression of DEN-2 virus titer (0.1 and 0.9 log10, respectively). P4-PMOs targeting the AUG translation start site region of the single open reading frame and the 5′ cyclization sequence region had moderate activity, generating 1.6- and 1.8-log10 reductions. Two P4-PMO compounds, 5′SL and 3′CS (targeting the 5′-terminal nucleotides and the 3′ cyclization sequence region, respectively), were highly efficacious, each reducing the viral titer by greater than 5.7 log10 compared to controls at 6 days after infection with DEN-2 virus. Further experiments showed that 5′SL and 3′CS inhibited DEN-2 virus replication in a dose-dependent and sequence-specific manner. Treatment with 10 µM 3′CS reduced the titers of all four DEN virus serotypes, i.e., DEN-1 (strain 16007), DEN-2 (16681), DEN-3 (16562), and DEN-4 (1036) viruses by over 4 log10, in most cases to below detectable limits. The extent of 3′CS efficacy was affected by the timing of compound application in relation to viral infection of the cells. The 5′SL and 3′CS P4-PMOs did not suppress the replication of West Nile virus NY99 in Vero cells. These data indicate that further evaluation of the 5′SL and 3′CS compounds as potential DEN virus therapeutics is warranted. [ABSTRACT FROM AUTHOR]

RNA elements within flavivirus genomes are potential targets for antiviral therapy. A panel of phosphorodiamidate morpholino oligomers (PMOs), whose sequences are complementary to RNA elements located in the 5′- and 3′-termini of the West Nile (WN) virus genome, were designed to anneal to important cis-acting elements and potentially to inhibit WN infection. A novel Arg-rich peptide was conjugated to each PMO for efficient cellular delivery. These PMOs exhibited various degrees of antiviral activity upon incubation with a WN virus luciferase-replicon-containing cell line. Among them, PMOs targeting the 5′-terminal 20 nucleotides (5′End) or targeting the 3′-terminal element involved in a potential genome cyclizing interaction (3′CSI) exhibited the greatest potency. When cells infected with an epidemic strain of WN virus were treated with the 5′End or 3′CSI PMO, virus titers were reduced by approximately 5 to 6 logs at a 5 µM concentration without apparent cytotoxicity. The 3′CSI PMO also inhibited mosquito-borne flaviviruses other than WN virus, and the antiviral potency correlated with the conservation of the targeted 3′CSI sequences of specific viruses. Mode-of-action analyses showed that the 5′End and 3′CSI PMOs suppressed viral infection through two distinct mechanisms. The 5′End PMO inhibited viral translation, whereas the 3′CSI PMO did not significantly affect viral translation but suppressed RNA replication. The results suggest that antisense PMO-mediated blocking of cis-acting elements of flavivirus genomes can potentially be developed into an anti-flavivirus therapy. In addition, we report that although a full-length WN virus containing a luciferase reporter (engineered at the 3′ untranslated region of the genome) is not stable, an early passage of this reporting virus can be used to screen for inhibitors against any step of the virus life cycle. [ABSTRACT FROM AUTHOR]

Although the Myc oncogene has long been known to play a role in many human cancers, the mechanisms that mediate its effects in both normal cells and cancer cells are not fully understood. We have initiated a genetic analysis of the Drosophila homolog of the Myc oncoprotein (dMyc), which is encoded by the dm locus. We carried out mosaic analysis to elucidate the functions of dMyc in the germline and somatic cells of the ovary during oogenesis, a process that involves cell proliferation, differentiation and growth. Germline and somatic follicle cells mutant for dm exhibit a profound decrease in their ability to grow and to carry out endoreplication, a modified cell cycle in which DNA replication occurs in the absence of cell division. In contrast to its dramatic effects on growth and endoreplication, dMyc is dispensable for the mitotic division cycles of both germline and somatic components of the ovary. Surprisingly, despite their impaired ability to endoreplicate, dm mutant follicle cells appeared to carry out chorion gene amplification normally. Furthermore, in germline cysts in which the dm mutant cells comprised only a subset of the 16-cell cluster, we observed strictly cell-autonomous growth defects. However, in cases in which the entire germline cyst or the whole follicular epithelium was mutant for dm, the growth of the entire follicle, including the wild-type cells, was delayed. This observation indicates the existence of a signaling mechanism that acts to coordinate the growth rates of the germline and somatic components of the follicle. In summary, dMyc plays an essential role in promoting the rapid growth that must occur in both the germline and the surrounding follicle cells for oogenesis to proceed. [ABSTRACT FROM AUTHOR]

Androgen deprivation therapy causes a paradoxical elevation of matrix metalloproteinases (MMPs) including MMP-9 resulting in aggressive tumor phenotype in many patients with prostate cancer. In this study, we have evaluated a novel antisense phosphorodiamidate Morpholino oligomer (PMO) targeted against MMP-9 in models of angiogenesis and in human prostate xenograft in athymic mice. The treatment of androgen-independent DU145 human prostate cells with a 21-mer MMP-9 antisense PMO caused a dose-dependent inhibition of cell proliferation compared to scrambled or MMP-2 antisense PMO at similar concentrations. This was associated with decreases in MMP-9 expression, gelatinolytic activity and increased stability of the insulin-like growth factor-binding protein (IGFBP-3), a proapoptotic factor and MMP-9 substrate. In vitro invasion assays revealed a 40-60% inhibition of DU145 cell invasion in the presence of 25?µM MMP-9 antisense PMO. A significant decrease in endothelial cell migration and vascularization was observed in the Matrigel plug assay in mice when treated intraperitoneally with 300?µg/day MMP-9 antisense for 21 days. In the highly vascular DU145 tumor xenografts, MMP-9 inhibition caused decreased tumor growth with regression in 50% of the animals. Histological analysis revealed increased apoptosis and fibrous tissue deposits in the MMP-9 antisense-treated tumors compared to the scrambled and saline controls. No apparent toxicity or mortality was associated with the MMP-9 PMO treatment. In summary, the MMP-9 antisense PMO inhibited in vitro prostate cancer cell proliferation, invasion and in vivo angiogenesis. These data establish the feasibility of developing a site-directed, nontoxic antisense therapeutic agent for inhibiting local invasion and metastasis.Cancer Gene Therapy (2003) 10, 823-832. doi:10.1038/sj.cgt.7700642 [ABSTRACT FROM AUTHOR]

Here we report the cloning and functional characterization of eight members of the UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase gene family from Drosophila melanogaster (polypeptide GalNAc transferase = pgant1-8). Full-length cDNAs were isolated from a Drosophila embryonic library based on homology to known ppGaNTases. Alignments with characterized mammalian isoforms revealed strong sequence similarities between certain fly and mammalian isoforms, highlighting putative orthologues between the species. In vitro activity assays demonstrated biochemical transferase activity for each gene, with three isoforms requiring glycosylated substrates. Comparison of the activities of Drosophila and mammalian orthologues revealed conservation of substrate preferences against a panel of peptide and glycopeptide substrates. Furthermore, Edman degradation analysis demonstrated that preferred sites of GalNac addition were also conserved between certain fly and mammalian orthologues. Semiquantitative PCR amplification of Drosophila cDNA revealed expression of most isoforms at each developmental stage, with some isoforms being less abundant at certain stages relative to others. In situ hybridization to Drosophila embryos revealed specific staining of pgant5 and pgant6 in the salivary glands and pgant5 in the developing hindgut. Additionally, pgant5 and pgant6 expression within the egg chamber was restricted to the follicle cells, cells known to be involved in egg formation and subsequent embryonic patterning. The characterization reported here provides additional insight into the use of this model system to dissect the biological role of this enzyme family in vivo during both fly and maremalian development. [ABSTRACT FROM AUTHOR]

The development of the head and tail regions of the Drosophila embryo is dependent upon the localized polar activation of Torso (Tor), a receptor tyrosine kinase that is uniformly distributed in the membrane of the developing embryo . Trunk (Trk), the proposed ligand for Tor, is secreted as an inactive precursor into the perivitelline fluid that lies between the embryonic membrane and the vitelline membrane (VM), the inner layer of the eggshell . The spatial regulation of Trk processing is thought to be mediated by the secreted product of the torsolike (tsl) gene, which is expressed during oogenesis by a specialized population of follicle cells present at the two ends of the oocyte . We show here that Tsl protein is specifically localized to the polar regions of the VM in laid eggs. We further demonstrate that although Tsl can associate with nonpolar regions of the VM, the activity of polar-localized Tsl is enhanced, suggesting the existence of another spatially restricted factor acting in this pathway. The incorporation of Tsl into the VM provides a mechanism for the transfer of spatial information from the follicle cells to the developing embryo. To our knowledge, Tsl represents the first example of an embryonic patterning determinant that is a component of the eggshell. [Copyright &y& Elsevier]

Backgroud:Neointimal hyperplasia following percutaneous transluminal coronary angioplasty (PTCA) is one of the major components of the process of restenosis. We evaluated the long-term impact of local delivery of c-myc neutrally charged antisense oligonucleotides (Resten-NG) upon neointimal formation following PTCA in a rabbit model.Methods:PTCA was performed in the iliac arteries of 10 New Zealand white rabbits at 8 atm for 30 s, three times. An infusion of 500 μg Resten-NG (n=6) or saline (n=4) was delivered to the site at 2 atm via the outer balloon pores of the transport™ catheter over 2 min. The diet was supplemented with 0.25% cholesterol for 10 days before and 6 months following PTCA.Results:After 6 months, animals were sacrificed and vessels were fixed in formalin, processed and stained with hematoxylin, eosin, and movat. Histological analysis revealed complete vascular healing in both groups of animals. Planimetry showed that intimal areas were 1.71±0.25 and 0.65±0.36 mm2 in the control and antisense delivery groups, respectively (P<.05).Conclusion:We conclude that local delivery of Resten-NG significantly inhibited neointimal thickening following PTCA in a rabbit for up to 6 months. [Copyright &y& Elsevier]

Recent studies in vertebrates and Drosophila melanogaster have revealed that Fringe-mediated activation of the Notch pathway has a role in patterning cell layers during organogenesis. In these processes, a homeobox-containing transcription factor is responsible for spatially regulating fringe (fng) expression and thus directing activation of the Notch pathway along the fng expression border. Here we show that this may be a general mechanism for patterning epithelial cell layers. At three stages in Drosophila oogenesis, mirror (mirr) and fng have complementary expression patterns in the follicle-cell epithelial layer, and at all three stages loss of mirr enlarges, and ectopic expression of mirr restricts, fng expression, with consequences for follicle-cell patterning. These morphological changes are similar to those caused by Notch mutations. Ectopic expression of mirr in the posterior follicle cells induces a stripe of rhomboid (rho) expression and represses pipe (pip), a gene with a role in the establishment of the dorsal-ventral axis, at a distance. Ectopic Notch activation has a similar long-range effect on pip. Our results suggest that Mirror and Notch induce secretion of diffusible morphogens and we have identified TGF-β (encoded by dpp) as such a molecule in germarium. We also found that mirr expression in dorsal follicle cells is induced by the EGF-receptor (EGFR) pathway and that mirr then represses pip expression in all but the ventral follicle cells, connecting EGFR activation in the dorsal follicle cells to repression of pip in the dorsal and lateral follicle cells. Our results suggest that the differentiation of ventral follicle cells is not a direct consequence of germline signalling, but depends on long-range signals from dorsal follicle cells, and provide a link between early and late events in Drosophila embryonic dorsal-ventral axis formation. [ABSTRACT FROM AUTHOR]

Presents abstracts of papers from the Medical Scientist Training Symposium scheduled on October 18-19, 1998 in Mohonk, New York City. 'Cellular Immune Response to Human Immunodeficiency Virus Infection'; 'A Kaptivating Protein: Intracellular Targeting of Hormonal Signals'; 'Characterization of the Mechanism of Transcriptional Activation by the Sea Urchin Transcription Factor, SSAP.'

Mitochondria have emerged as important determinants in cancer progression and malignancy. However, the role of mitochondrial membranes in cancer onset and progression has not been thoroughly investigated. This study compares the structural and functional properties of mitochondrial membranes in prostate and colon cancer cells in comparison to normal mitochondria, and possible therapeutic implications of these membrane changes. Specifically, isolation of cell mitochondria and preparation of inverted sub-mitochondrial particles (SMPs) illuminated significant cancer-induced modulations of membrane lipid compositions, fluidity, and activity of cytochrome c oxidase, one of the key mitochondrial enzymes. The experimental data further show that cancer-associated membrane transformations may account for mitochondria targeting by betulinic acid and resveratrol, known anti-cancer molecules. Overall, this study probes the relationship between cancer and mitochondrial membrane transformations, underlying a potential therapeutic significance for mitochondrial membrane targeting in cancer. Unlabelled Image • Transformation of lipid composition in colon and prostate cancers identified • Significantly increased concentration of cardiolipin detected • Elevated cardiolipin concentration was shown to affect enzyme functionalities in sub-mitochondria particles (SMPs) • The elevated cardiolipin level in cancer mitochondria is linked with mitochondrial targeting by betulinic acid and reservatrol. [ABSTRACT FROM AUTHOR]

Background: Many enteroviruses as well as hepatitis C have been increasingly found in association with cardiomyopathies, even though, the evidence has been inconclusive. Methods: From 1998 to 2001, at the liver clinic of our community-based hospital, in the cohort of hepatitis C patients, infection evidence by PCR confirmation and echocardiographic measures of left ventricular (LV) function were obtained. Results: One hundred-eight patients were identified. Fifty percent of them have echocardiographic abnormalities. Patient''s mean age was 55±10 years; 45% women, 55% men; 32% had LVH, 13.8% had high right ventricular systolic pressures, and 4% displayed contractility abnormalities. No cases of dilated or hypertrophic cardiomiopathies were found. Conclusions: Echocardiographic abnormalities are not uncommon findings associated with hepatitis C infection, however all these findings can easily be explained by existing co-morbidities. High right ventricular systolic pressure, suggestive of pulmonary hypertension, which may be secondary to portal hypertension, is an independent finding for hepatitis C. [Copyright &y& Elsevier]

Background As minimally invasive spine surgery evolves, spine surgeons increasingly rely on advanced intraoperative computed tomography (iCT). iCT provides rapid acquisition of high-resolution images, reduces radiation exposure, improves surgical accuracy, and decreases operative time. However, all iCT systems currently available pose a patient safety risk as their physical space requirements limit patient access in the event of an emergency, particularly when patients are in the prone position. After a near-cardiac arrest at our institution during posterior cervical spine surgery, it was apparent that the presence of the iCT complicated the ability to rapidly reposition the patient in order to provide appropriate resuscitation. Methods To ensure our ability to provide timely care during an emergency, we determined that a process which included all members of the operating room (OR) team was required. We held an initial planning meeting where a detailed plan-of-action was created, reviewed, and revised in response to feedback from all stakeholders. We then simulated a cardiac arrest to test our resuscitation plan with all members of the neurosurgery team. A mannequin was positioned prone on an OR table within the iCT, and a resuscitation plan was created. Results The team orchestrated the mock resuscitation, and the time of cardiac arrest in the prone position to supine repositioning required 110 seconds. The simulation was recorded for post-"code" performance review. Application of the protocol during an actual cardiac arrest was associated with successful restoration of spontaneous circulation and full recovery. Conclusions The development and rehearsal of an emergency plan of action greatly facilitated the timely responsiveness of the neurosurgical OR team during a simulated cardiac arrest and was an effective way to identify and address key logistical issues regarding the use of an iCT system. Highlights • To ensure timely, effective care during an IA emergency when iCT is used, we determined that a process. • All members of the operating room (OR) team were involved. • A detailed plan was created, reviewed, and revised in response to feedback from all neurosurgical OR team members. • We simulated a cardiac arrest to test our resuscitation plan with all members of the neurosurgery OR team participating. • The plan enabled us to reposition a "patient" in head pins on an iCT table from prone to supine position in 110 seconds. [ABSTRACT FROM AUTHOR]

Spatially regulated transcription of the pipe gene in ventral cells of the Drosophila ovary follicle cell epithelium is a key event that specifies progeny embryo dorsal-ventral (DV) polarity. pipe encodes ten putative protein isoforms, all of which exhibit similarity to vertebrate glycosaminoglycan-modifying enzymes. Expression of one of the isoforms, Pipe-ST2, in follicle cells has previously been shown to be essential for DV patterning. pipe is also expressed in the embryonic salivary gland and its expression there is required for normal viability. Here, we show that in addition to Pipe-ST2, seven of the other Pipe isoforms are expressed in the ovary, whereas all Pipe isoforms are abundantly expressed in the embryo. Of the ten isoforms, only Pipe-ST2 can restore ventral and lateral pattern elements to the progeny of otherwise pipe-null mutant females. By contrast, three Pipe isoforms, but not Pipe-ST2, support the production of a novel pipe-dependent epitope present in the embryonic salivary gland. These data indicate that differences in functional specificity, and presumably enzymatic specificity, are associated with several of the Pipe isoforms. In addition, we show that uniform expression of the Pipe-ST2 isoform in the follicle cell layer of females otherwise lacking pipe expression leads to the formation of embryos with a DV axis that is appropriately oriented with respect to the intrinsic polarity of the eggshell. This suggests the existence of a second mechanism that polarizes the Drosophila embryo, in addition to the ventrally restricted transcription of the pipe gene. [ABSTRACT FROM AUTHOR]

Objectives: To determine the antiviral activity of phosphorodiamidate morpholino oligomers (PMO) and peptide-conjugated PMO (PPMO) in AG129 mice infected with dengue 2 virus (DENV-2).Methods: Antisense PMO and PPMO were designed against the 5' terminal region (5'SL) or the 3'-cyclization sequence region (3'CS) of DENV genomic RNA and administered to AG129 mice before and/or after infection with DENV-2. In addition, cell culture evaluations designed to determine optimum PPMO length, and pharmacokinetic and toxicity analysis of PPMO were also carried out.Results: Mock-treated AG129 mice lived for 9-17 days following intraperitoneal (ip) infection with 10(4)-10(6) pfu of DENV-2 (strain New Guinea C). Intraperitoneal administration of 5'SL or 3'CS PPMO before and after DENV infection produced an increase in the average survival time of up to 8 days. Animals receiving only post-infection PPMO treatment did not benefit significantly. Cell culture studies showed that PPMO of 22-24 bases long produced substantially higher DENV titre reductions than did PPMO that were either shorter or longer. Pharmacokinetic and toxicology analysis with non-infected animals showed that nine consecutive once-daily ip treatments of 10 mg/kg PPMO resulted in high concentrations of PPMO in the liver and caused little impact on overall health.Conclusions: The data indicate that PPMO had considerable antiviral efficacy against DENV-2 in the AG129 mouse model and that PPMO treatment early in the course of an infection was critical to extending the survival times of DENV-2-infected mice in the AG129 model system. [ABSTRACT FROM AUTHOR]

The recent emergence of novel pathogenic human and animal coronaviruses has highlighted the need for antiviral therapies that are effective against a spectrum of these viruses. We have used several strains of murine hepatitis virus (MHV) in cell culture and in vivo in mouse models to investigate the antiviral characteristics of peptide-conjugated antisense phosphorodiamidate morpholino oligomers (P-PMOs). Ten P-PMOs directed against various target sites in the viral genome were tested in cell culture, and one of these (5TERM), which was complementary to the 5′ terminus of the genomic RNA, was effective against six strains of MHV. Further studies were carried out with various arginine-rich peptides conjugated to the 5TERM PMO sequence in order to evaluate efficacy and toxicity and thereby select candidates for in vivo testing. In uninfected mice, prolonged P-PMO treatment did not result in weight loss or detectable histopathologic changes. 5TERM P-PMO treatment reduced viral titers in target organs and protected mice against virus-induced tissue damage. Prophylactic 5TERM P-PMO treatment decreased the amount of weight loss associated with infection under most experimental conditions. Treatment also prolonged survival in two lethal challenge models. In some cases of high-dose viral inoculation followed by delayed treatment, 5TERM P-PMO treatment was not protective and increased morbidity in the treated group, suggesting that P-PMO may cause toxic effects in diseased mice that were not apparent in the uninfected animals. However, the strong antiviral effect observed suggests that with further development, P-PMO may provide an effective therapeutic approach against a broad range of coronavirus infections. [ABSTRACT FROM AUTHOR]