Locoregional therapy of locally advanced breast cancer: a clinical practice guideline.
In female patients with locally advanced breast cancer (labc) and good response to neoadjuvant chemotherapy (nact), including endocrine therapy, what is the role of breast-conserving surgery (bcs) compared with mastectomy?In female patients with labc, is radiotherapy (rt) indicated for those who have undergone mastectomy?does locoregional rt, compared with breast or chest wall rt alone, result in a higher survival rate and lower recurrence rates?is rt indicated for those achieving a pathologic complete response (pcr) to nact?In female patients with labc who receive nact, is the most appropriate axillary staging procedure sentinel lymph node biopsy (slnb) or axillary dissection? Is slnb indicated before nact rather than at the time of surgery?How should female patients with labc that does not respond to initial nact be treated? This guideline was developed by Cancer Care Ontario's Program in Evidence-Based Care (pebc) and the Breast Cancer Disease Site Group (dsg). A systematic review was prepared based on literature searches conducted using the medline and embase databases for the period 1996 to December 11, 2013. Guidelines were located from that search and from the Web sites of major guideline organizations. The working group drafted recommendations based on the systemic review. The systematic review and recommendations were then circulated to the Breast Cancer dsg and the pebc Report Approval Panel for internal review; the revised document underwent external review. The full three-part evidence series can be found on the Cancer Care Ontario Web site. For most patients with labc, modified radical mastectomy should be considered the standard of care. For some patients with noninflammatory labc, bcs can be considered on a case-by-case basis when the surgeon deems that the disease can be fully resected and the patient expresses a strong preference for breast preservation.For patients with labc, rt after mastectomy is recommended.It is recommended that, after bcs or mastectomy, patients with labc receive locoregional rt encompassing the breast or chest wall and local node-bearing areas.It is recommended that postoperative rt remain the standard of care for patients with labc who achieve pcr to nact.It is recommended that axillary dissection remain the standard of care for axillary staging in labc, with the judicious use of slnb in patients who are advised of the limitations of the current data.Although slnb either before or after nact is technically feasible, the data are insufficient to make any recommendation about the optimal timing of slnb with respect to nact. Limited data suggest higher sentinel lymph node identification rates and lower false negative identification rates when slnb is conducted before nact; however, those data must be balanced against the requirement for two operations if slnb is not performed at the time of resection of the main tumour.It is recommended that patients receiving neoadjuvant anthracycline-taxane-based therapy (or other sequential regimens) whose tumours do not respond to the initial agent or agents, or who experience disease progression, be expedited to the next agent or agents of the regimen.For patients who, in the opinion of the treating physician, fail to respond or progress on first-line nact, several therapeutic options can be considered, including second-line chemotherapy, hormonal therapy (if appropriate), rt, or immediate surgery (if technically feasible). Treatment should be individualized through discussion at a multidisciplinary case conference, considering tumour characteristics, patient factors and preferences, and risk of adverse effects.It is recommended that prospective randomized clinical trials be designed for patients with labc who fail to respond to nact so that more definitive treatment recommendations can be developed.


[Eyelid tumor in anophthalmic socket due to retinoblastoma: report of two cases].
Retinoblastoma is a unilateral or bilateral intraocular tumor, occurring as a hereditary or sporadic tumor. Another not ocular lesion is a possibility for retinoblastoma carriers. The authors report the cases of two patients: a son with bilateral anophthalmic socket due to retinoblastoma, who presented unilateral upper eyelid tumor; and his father with unilateral anophthalmic socket secondary to enucleation due to retinoblastoma and a tumor growing fast in the upper eyelid at the same side of the anophthalmic cavity. Lesions were diagnosed as sebaceous carcinoma confirmed by immunohistochemical exam. Both patients had the eyelid tumors removed, with good resolution. None of them have presented either signs of recurrence or metastasis.


An Introduction to Probabilistic Record Linkage with a Focus on Linkage Processing for WTC Registries.
Since its post-World War II inception, the science of record linkage has grown exponentially and is used across industrial, governmental, and academic agencies. The academic fields that rely on record linkage are diverse, ranging from history to public health to demography. In this paper, we introduce the different types of data linkage and give a historical context to their development. We then introduce the three types of underlying models for probabilistic record linkage: Fellegi-Sunter-based methods, machine learning methods, and Bayesian methods. Practical considerations, such as data standardization and privacy concerns, are then discussed. Finally, recommendations are given for organizations developing or maintaining record linkage programs, with an emphasis on organizations measuring long-term complications of disasters, such as 9/11.


Cleavage-stage biopsy significantly impairs human embryonic implantation potential while blastocyst biopsy does not: a randomized and paired clinical trial.
To determine if cleavage- or blastocyst-stage embryo biopsy affects reproductive competence. Paired randomized clinical trial. Academic-assisted reproduction program. Attempting conception through IVF. After selecting two embryos for transfer, one was randomized to biopsy and the other to control. Both were transferred within shortly thereafter. The biopsy was submitted for microarray analysis and single-nucleotide polymorphism (SNP) profiling. Buccal DNA obtained from the neonate after delivery had microarray analysis and SNP profile compared with that of the embryonic DNA. A match confirmed that the biopsied embryo implanted and developed to term, whereas a nonmatch indicated that the control embryo had led to the delivery. Paired analysis of the delivery rates of the transferred embryos. Either twin delivery or failure to deliver represents equivalent outcomes for the biopsied and control embryos. In contrast, singletons were determined to be from the biopsied or the control embryo. Blastomere biopsy on day 3 of development resulted in a significant reduction in sustained implantation. Only 30% of biopsied embryos had sustained implantation and ultimately developed into live-born infants versus 50% of unbiopsied controls, a relative reduction of 39%. In contrast, sustained implantation rates were equivalent (51% vs. 54%) for biopsied and control blastocysts. Cleavage-stage biopsy markedly reduced embryonic reproductive potential. In contrast, trophectoderm biopsy had no measurable impact and may be used safely when embryo biopsy is indicated. NCT01219504.


An international survey to identify the intrinsic and extrinsic factors of research studies most likely to change orthopaedic practice.
Evidence -based medicine (EBM) is designed to inform clinical decision-making within all medical specialties, including orthopaedic surgery. We recently published a pilot survey of the Canadian Orthopaedic Association (COA) membership and demonstrated that the adoption of EBM principles is variable among Canadian orthopaedic surgeons. The objective of this study was to conduct a broader international survey of orthopaedic surgeons to identify characteristics of research studies perceived as being most influential in informing clinical decision-making. A 29-question electronic survey was distributed to the readership of an established orthopaedic journal with international readership. The survey aimed to analyse the influence of both extrinsic (journal quality, investigator profiles, etc.) and intrinsic characteristics (study design, sample size, etc.) of research studies in relation to their influence on practice patterns. A total of 353 surgeons completed the survey. Surgeons achieved consensus on the 'importance' of three key designs on their practices: randomised controlled trials (94%), meta-analyses (75%) and systematic reviews (66%). The vast majority of respondents support the use of current evidence over historical clinical training; however subjective factors such as journal reputation (72%) and investigator profile (68%) continue to influence clinical decision-making strongly. Although intrinsic factors such as study design and sample size have some influence on clinical decision-making, surgeon respondents are equally influenced by extrinsic factors such as investigator reputation and perceived journal quality.Cite this article: Dr M. Ghert. An international survey to identify the intrinsic and extrinsic factors of research studies most likely to change orthopaedic practice. Bone Joint Res 2016;5:130-136. DOI: 10.1302/2046-3758.54.2000578.


A decentralized control of interconnected systems using neural networks.
We develop a decentralized neural-network (NN) controller for a class of large-scale nonlinear systems with the high-order interconnections. The controller is a mixed NN comprised of a conventional NN and a special NN. The conventional NN is used to approximate the unknown nonlinearities in the subsystem, while a special NN is used to counter the high-order interconnections. We prove that this NN structure can achieve a stable controller for the large-scale systems.


Uncovering the asymmetric impacts of economic policy uncertainty on green financial markets in China.
Green finance is considered a novel tool of financing to promote the development of the green economy, which has huge investment attractiveness. Previous studies detected the impacts of economic policy uncertainty (EPU) on the green financial markets are mixed. To this end, this study deeply investigates the asymmetric and heterogeneous impacts of EPU on the green bond and green stock markets based on the quantile-on-quantile (QQ) method. Using the data of China Economic Policy Uncertainty Index (CNEPU) and green financial indices, we get some interesting results. (1) EPU has an overall negative effect on the green financial markets, and the green stock market reacts more strongly than the green bond market. (2) For green bond market, the higher quantiles of EPU on a bear market have more significant effect than that on a bull market. (3) For green stock market, a negative effect of EPU on green stock market is observed for the lower quantiles of EPU, while a positive effect is noted at the highest quantiles of EPU. This paper could provide a reference for investors making green investment strategies and for policymakers making policies to promote green development.


The safety and efficacy of intra-articular dual molecular weighted hyaluronic Acid in the treatment of knee osteoarthritis: the I.d.e.h.a. Study.
In clinical practice viscosupplementation with hyaluronic acid (HA) is common for the treatment of degenerative osteoarthritis (OA). Both molecular weight and concentration of HA have significant impact on its rheological properties, which in turn affects its therapeutic effects. The objective of this study is to evaluate the effectiveness of a double HA preparation for the treatment of knee osteoarthritis with respect to pain reduction, joint function improvement and concomitant medication consumption reduction. One thousand and fourteen patients (521 males and 693 females) with a mean age of 62.4 years old, suffering from OA of the knee, were enrolled into this study. All patients received two intra-articular injections one week apart and a third injection one month after the second one. Concomitant medication was recorded and evaluated at follow up visits. Evaluation was performed at baseline, day 30 and day 180, on several parameters: knee pain by visual analog scale (VAS) 0-10 cm, Lequesne Index, and consumption of concomitant medications including non-steroidal anti-inflammatory drugs, analgesics and chondoprotective supplementations. A statistically significant reduction in pain VAS score was recorded at D30 (38.01±17.68; P<0.01) before the third injection, and D180 (25.91±15.33; P<0.01) check-points comparing to baseline (67.12±15.99). Similarly, remarkable reduction in Lequesne Index was shown at D30 (5.91±4.01; P<0.01) in 1214 patients before the third injection, and D180 (3.59±3.45; P<0.01) (with 938 patients) when compared to the baseline (11.60±5.13). Patients also consumed less concomitant medications after the treatment course. The beneficial effects were maintained for up to six months. Intra-articular injection of a double HA preparation of low molecular weight and high molecular weight of different concentrations was well tolerated, and generated satisfactory results in terms of pain control, joint function improvement and concomitant medication reduction for the management of knee OA.


[Organ-preserving operations in the treatment of retroampullary duodenal ulcers].
The authors treated 106 patients with retroampullary duodenal ulcer. Resections were performed in 16, the organ-preserving operations--in 89 patients. In 58 (64.8%) cases, the duodenoplasty with the different types of vagotomy and preservation of the pyloric sphincter using the techniques developed in the clinic was performed.


Glomerular filtration rate decline as a surrogate end point in kidney disease progression trials.
Chronic kidney disease (CKD) is strongly associated with increased risks of progression to end-stage kidney disease (ESKD) and mortality. Clinical trials evaluating CKD progression commonly use a composite end point of death, ESKD or serum creatinine doubling. However, due to low event rates, such trials require large sample sizes and long-term follow-up for adequate statistical power. As a result, very few interventions targeting CKD progression have been tested in randomized controlled trials. To overcome this problem, the National Kidney Foundation and Food and Drug Administration conducted a series of analyses to determine whether an end point of 30 or 40% decline in estimated glomerular filtration rate (eGFR) over 2-3 years can substitute for serum creatinine doubling in the composite end point. These analyses demonstrated that these alternate kidney end points were significantly associated with subsequent risks of ESKD and death. However, the association between, and consistency of treatment effects on eGFR decline and clinical end points were influenced by baseline eGFR, follow-up duration and acute hemodynamic effects. The investigators concluded that a 40% eGFR decline is broadly acceptable as a kidney end point across a wide baseline eGFR range and that a 30% eGFR decline may be acceptable in some situations. Although these alternate kidney end points could potentially allow investigators to conduct shorter duration clinical trials with smaller sample sizes thereby generating evidence to guide clinical decision-making in a timely manner, it is uncertain whether these end points will improve trial efficiency and feasibility. This review critically appraises the evidence, strengths and limitations pertaining to eGFR end points.


Genomic skimming and nanopore sequencing uncover cryptic hybridization in one of world's most threatened primates.
The Brazilian buffy-tufted-ear marmoset (Callithrix aurita), one of the world's most endangered primates, is threatened by anthropogenic hybridization with exotic, invasive marmoset species. As there are few genetic data available for C. aurita, we developed a PCR-free protocol with minimal technical requirements to rapidly generate genomic data with genomic skimming and portable nanopore sequencing. With this direct DNA sequencing approach, we successfully determined the complete mitogenome of a marmoset that we initially identified as C. aurita. The obtained nanopore-assembled sequence was highly concordant with a Sanger sequenced version of the same mitogenome. Phylogenetic analyses unexpectedly revealed that our specimen was a cryptic hybrid, with a C. aurita phenotype and C. penicillata mitogenome lineage. We also used publicly available mitogenome data to determine diversity estimates for C. aurita and three other marmoset species. Mitogenomics holds great potential to address deficiencies in genomic data for endangered, non-model species such as C. aurita. However, we discuss why mitogenomic approaches should be used in conjunction with other data for marmoset species identification. Finally, we discuss the utility and implications of our results and genomic skimming/nanopore approach for conservation and evolutionary studies of C. aurita and other marmosets.


The marrow colony forming cell and serum colony stimulating factor of the chicken.
Marrow cells of the chicken produced colonies in semisolid media. Developing colonies consisted of granulocytes, macrophages or a mixture of these two cell types. The granulocyte-macrophage CFC was nonadherent. An adherent 'CFC' was also present and it differed in several ways from the nonadherent CFC: (a) clones contained only macrophages, (b) they contained a core of nonrefractile cells, (c) their appearance was delayed 1-2 weeks, (d) they were unaffected by the presence of erythrocytes and (e) the efficiency of cloning was increased but the percentage of clones able to produce 50 or more cells was markedly decreased, i.e., the cluster/colony ratio was increased. The growth of both colony types was strictly dependent on the presence of CSF. Data obtained from dose-response studies on unfractionated marrow indicated that clusters and colonies were derived from single cells. The CSF of chicken serum yielded sigmoid dose-response curves when tested on marrow cells. Calf serum could not support cluster or colony formation when tested alone but it did have an enhancing effect on the CSF of chicken serum. Levels of serum CSF were increased by injecting chickens with bacterial endotoxin. This phenomenon occurred with five chicken lines tested, but certain chickens of the Kimber line did not respond to endotoxin with elevated levels of CSF.


The HNF1/HNF4-dependent We2 element of woodchuck hepatitis virus controls viral replication and can activate the N-myc2 promoter.
Transcriptional activation of myc family proto-oncogenes through the insertion of viral sequences is the predominant mechanism by which woodchuck hepatitis virus (WHV) induces liver tumors in chronically infected animals. The main target is N-myc2, a functional retroposon of the N-myc gene, but c-myc and N-myc are also marginally involved. Here we identify a major, liver-specific regulatory element in the WHV genome (We2) which efficiently activates the N-myc2 promoter in cultured hepatoma cells. In the context of the episomal viral genome, We2 governs the production of pregenomic RNA and thus plays a central role in the control of viral replication. We2 activity is primarily controlled by the liver-enriched HNF1 and HNF4 transcription factors, although NF1 and Oct proteins were also shown to bind in a central region. The expression of HNF1 and HNF4 appears to be maintained in woodchuck tumors. Thus, We2 is a prime candidate for controlling myc gene cis activation during WHV-induced hepatocarcinogenesis.


Outer membrane proteins for serologic detection of Ornithobacterium rhinotracheale infection in turkeys.
Ornithobacterium rhinotracheale (ORT) is a bacterium responsible for a respiratory disease in turkeys and chickens and has been identified as one of the emerging respiratory bacterial pathogens. The clinical signs and lesions caused by ORT are very similar to those caused by other respiratory infectious agents; therefore, an accurate diagnostic test is necessary to identify the infection. In this study, we have investigated the use of outer membrane proteins of ORT in an indirect enzyme-linked immunosorbent assay (ELISA) to detect the exposure to ORT infection. Outer membrane proteins of ORT were extracted and used as an antigen in ELISA to detect infection in turkeys exposed to different serotypes of ORT. The ELISA results were compared with the conventional serum plate agglutination test. The agglutination test detected specific antibodies for ORT in 65% of experimentally infected turkeys during the first 2 wk of infection. The ELISA detected up to 100% of infected birds for 8 wk postinfection. The results suggest that ELISA is able to detect the exposure to ORT in later stages of the infection and this assay can be used in serologic surveillance of ORT infection for poultry in the field.


Fluorescent Multiplex Cell Rolling Assay: Simultaneous Capturing up to Seven Samples in Real-Time Using Spectral Confocal Microscopy.
The parallel plate flow chamber assay is widely utilized to study physiological cell-cell adhesive interactions under dynamic flow that mimics the bloodstream. In this technique, the cells are perfused under defined shear stresses over a monolayer of endothelial cells (expressing homing molecules, e.g., selectins) or a surface (expressing recombinant homing molecules). However, with the need to study multiple samples and multiple parameters per sample, using a traditional bright-field microscope-based flow assay allows only one sample at a time to be analyzed, resulting in high interexperiment variability, the need for normalization, waste of materials, and significant consumption of time. We developed a multiplexing approach using a three-color fluorescence staining method, which allowed for up to seven different combination signatures to be run at one time. Using this fluorescent multiplex cell rolling (FMCR) assay, each sample is labeled with a different signature of emission wavelengths and mixed with other samples just minutes before the flow run. Subsequently, real-time images are acquired in a single pass using a line-scanning spectral confocal microscope. To illustrate the glycan-dependent binding of E-selectin, a central molecule in cell migration, to its glycosylated ligands expressed on myeloid-leukemic cells in flow, the FMCR assay was used to analyze E-selectin-ligand interactions following the addition (fucosyltransferase-treatment) or removal (deglycosylation) of key glycans on the flowing cells. The FMCR assay allowed us to analyze the cell-adhesion events from these different treatment conditions simultaneously in a competitive manner and to calculate differences in rolling frequency, velocity, and tethering capability of cells under study.


Direct imaging of photonic nanojets.
We report the direct experimental observation of photonic nanojets created by single latex microspheres illuminated by a plane wave at a wavelength of 520 nm. Measurements are performed with a fast scanning confocal microscope in detection mode, where the detection pinhole defines a diffraction-limited observation volume that is scanned in three dimensions over the microsphere vicinity. From the collected stack of images, we reconstruct the full 3 dimensional photonic nanojet beam. Observations are conducted for polystyrene spheres of 1, 3 and 5 microm diameter deposited on a glass substrate, the upper medium being air or water. Experimental results are compared to calculations performed using the Mie theory. We measure nanojet sizes as small as 270 nm FWHM for a 3 microm sphere at a wavelength lambda of 520 nm. The beam keeps a subwavelength FWHM over a propagation distance of more than 3 lambda, displaying all the specificities of a photonic nanojet.


Protective action of sulfur compounds against aldehyde toxicants of cigarette smoke.
Acetaldehyde, acrolein, and formaldehyde are important toxicants of cigarette smoke. Since acetaldehyde is a toxicant common to both heavy alcohol drinking and heavy cigarette smoking, protection studies were undertaken in rats first against acetaldehyde lethality and later against acrolein and formaldehyde lethality. This paper summarizes findings from previously published studies. With acetaldehyde (LD 90 dose/72 hrs.) protection studies were carried out by pretreatment with a high oral dose (2 mmoles/kg) of L-cysteine and 17 other sulfur compounds. Good protection (80% or more survivors) was obtained with L-cysteine, N-acetylcysteine, thiamin . HCl, sodium metabisulfite, L-cysteic acid, and a combination (CTA) of L-cysteine, thiamin . HCl, and L-ascorbic acid. N-acetylcysteine, L-cysteic acid, and the combination CTA gave 100% protection (i.e. 100% survivors). With acrolein (LD 95 dose/72 hrs.), protection data showed 80% survivors with L-cysteine and 95% with the combination CTA. With formaldehyde (LD 95 dose/72 hrs.), protection resulted in 20% survivors with L-cysteine and 55% with the combination CTA. However, protection against formaldehyde lethality could be increased to 90% survivors by repeated dosing with L-ascorbic acid per se over a two day period prior to administration of formaldehyde. Further animal experimentation is necessary before any extrapolation of these findings for human use can be considered. Due to current interest, the potential value of N-acetylcysteine as a protectant is also discussed.


Solirubrobacter ginsenosidimutans sp. nov., isolated from soil of a ginseng field.
A Gram-reaction-positive, non-motile, non-spore-forming, aerobic rod, designated BXN5-15(T), was isolated from the soil of a ginseng field on Baekdu Mountain in China. Strain BXN5-15(T) grew optimally at 30 °C and pH 6.0-7.0 in the absence of NaCl on R2A agar. Strain BXN5-15(T) displayed β-glucosidase activity, which allowed it to transform ginsenoside Rb(1) (one of the dominant active components of ginseng) via Rd to minor ginsenoside F(2). On the basis of 16S rRNA gene sequence analysis, strain BXN5-15(T) was shown to belong to the genus Solirubrobacter. The closest phylogenetic relatives were Solirubrobacter soli Gsoil 355(T) (98.4 % 16S rRNA gene sequence similarity) and Solirubrobacter pauli B33D1(T) (96.4 %). Lower sequence similarities (<96.0 %) were found with all of the other recognized members of the order Solirubrobacterales. The predominant quinone was MK-7(H(4)). The major fatty acids (>10 %) were C(18 : 1)ω9c, iso-C(16 : 0) and C(18 : 3)ω6,9,12c. The G+C content of the genomic DNA was 70.6 mol%. DNA-DNA relatedness between strain BXN5-15(T) and S. soli KCTC 12628(T) was 23.3 %. On the basis of genotypic, phenotypic and chemotaxonomic data, strain BXN5-15(T) represents a novel species within the genus Solirubrobacter, for which the name Solirubrobacter ginsenosidimutans sp. nov. is proposed. The type strain is BXN5-15(T) ( = KACC 20671(T) = LMG 24459(T)).


New Guinean Issidae: description of new taxa in a poorly known island fauna (Hemiptera, Fulgoroidea).
In the framework of the recent expedition results of "Our Planet Reviewed Papua-New-Guinea 2012-2013" we provide here the first Issidae (Hemiptera, Fulgoroidea) fauna review of New Guinea with the description of three new taxa: one new genus Papunega Gnezdilov et Bourgoin gen. nov. with two new species: Papunega magnifacies Gnezdilov et Le Cesne, sp. nov. (type species), Papunega armocula Gnezdilov et Soulier-Perkins, sp. nov. Tetrica fasciatifrons Melichar, 1906 is transferred to the genus Papugena gen. nov. to become Papugena fasciatifrons (Melichar, 1906) comb. nov. New Guinean Issidae fauna now includes 7 genera and 16 species.


Active Site Dynamical Effects in the Hydrogen Transfer Rate-limiting Step in the Catalysis of Linoleic Acid by Soybean Lipoxygenase-1 (SLO-1): Primary and Secondary Isotope Contributions.
Using ab initio molecular dynamics (AIMD) simulations that facilitate the treatment of rare events, we probe the active site participation in the rate-determining hydrogen transfer step in the catalytic oxidation of linoleic acid by soybean lipoxygenase-1 (SLO-1). The role of two different active site components is probed. (a) On the hydrogen atom acceptor side of the active site, the hydrogen bonding propensity between the acceptor side hydroxyl group, which is bound to the iron cofactor, and the backbone carboxyl group of isoleucine (residue number 839) is studied toward its role in promoting the hydrogen transfer event. Primary and secondary (H/D) isotope effects are also probed and a definite correlation with subtle secondary H/D isotope effects is found. With increasing average nuclear kinetic energy, the increase in transfer probability is enhanced due to the presence of the hydrogen bond between the backbone carbonyl of I839 and the acceptor oxygen. Further increase in average nuclear kinetic energy reduces the strength of this secondary hydrogen bond which leads to a deterioration in hydrogen transfer rates and finally embrances an Arrhenius-like behavior. (b) On the hydrogen atom donor side, the coupling between vibrational modes predominantly localized on the donor-side linoleic acid group and the reactive mode is probed. There appears to be a qualitative difference in the coupling between modes that belong to linoleic acid and the hydrogen transfer mode, for hydrogen and deuterium transfer. For example, the donor side secondary hydrogen atom is much more labile (by nearly a factor of 5) during deuterium transfer as compared to the case for hydrogen transfer. This appears to indicate a greater coupling between the modes belonging to the linoleic acid scaffold and the deuterium transfer mode and also provides a new rationalization for the abnormal (nonclassical) secondary isotope effect results obtained by Knapp, Rickert, and Klinman in J. Am. Chem. Soc. , 2002 , 124 , 3865 . To substantiate our findings noted in point a above, we have suggested an I839 → A839 or I839 → V839 mutation. This will modify the bulkiness of hydrogen the bonding residue, allowing greater flexibility in the secondary hydrogen bond formation highlighted above and adversely affecting the reaction rate.


Expression of adrenomedullin and peptide amidation activity in human prostate cancer and in human prostate cancer cell lines.
After therapeutic hormone deprivation, prostate cancer (CaP) cells often develop androgen-independent growth through not-well-defined mechanisms. The presence of neuroendocrine (NE) cells is often greater in prostate carcinoma than in normal prostate, and the frequency of NE cells correlates with tumor malignancy, loss of androgen sensitivity, increase of autocrine-paracrine activity, and poor prognosis. In some CaPs, neuropeptides have been previously implicated as growth factors. Peptidylglycine alpha-amidating monooxygenase (PAM) is the enzyme producing alpha-amidated bioactive peptides from their inactive glycine-extended precursors. In the present work, we demonstrate that androgen-independent PC-3 and DU145 cell lines, derived from human CaP, express PAM in vitro and in xenografts implanted in athymic nude mice, indicating that they are able to produce alpha-amidated peptides. Contrarily, barely detectable levels of PAM were found in the androgen-sensitive LNCaP cell line. We also show that whereas PC-3 and DU145 cells produce and secrete adrenomedullin (AM), a multifunctional amidated peptide, no expression was found in LNCaP cells. We further demonstrate that AM acts as a growth factor for DU145 cells, which suggests the existence of an autocrine loop mechanism that could potentially drive neoplastic growth. PAM mRNA levels were found to be 3-fold higher in prostate adenocarcinomas compared with that of human benign prostate hyperplasia (BPH) as demonstrated by real-time quantitative reverse transcription-PCR. The analysis of AM message expression in BPH and CaP (Gleason's score, 6-9) shows a clear distinction between benign and CaP. The expression was detected only in adenocarcinomas tissues with a marked increase in samples with a high Gleason's score. Immunocytochemically, AM was localized in the carcinomatous epithelial compartment. NE phenotype, assessed after the immunocytochemical localization of neuron-specific enolase (NSE), was found in both the epithelial and the stromal compartments of cancers; in BPH, only some spare basal cells were NSE-labeled. Cancer progression could be accelerated by peptides secreted by a population of cells capable of inducing androgen-independent tumoral growth via autocrine-paracrine mechanisms.


Trailing-edge fringes enable robust aerodynamic force production and noise suppression in an owl wing model.
As one of the unique owl-wing morphologies, trailing-edge (TE) fringes are believed to play a critical role in the silent flight of owls and have been widely investigated using idealized single/tandem airfoils. However, the effect of TE fringes and associated mechanisms on the aeroacoustics of owl wings, which feature curved leading edges, wavy TEs, and several feather slots at the wingtips, have not yet been addressed. In this study, we constructed two 3D owl wing models, one with and one without TE fringes, based on the geometric characteristics of a real owl wing. Large-eddy simulations and the Ffowcs Williams‒Hawkings analogy were combined to resolve the aeroacoustic characteristics of the wing models. Comparisons of the computed aerodynamic forces and far-field acoustic pressure levels demonstrate that the fringes on owl wings can robustly suppress aerodynamic noise while sustaining aerodynamic performance comparable to that of a clean wing. By visualizing the near-field flow dynamics in terms of flow and vortex structures as well as flow fluctuations, the mechanisms of TE fringes in owl wing models are revealed. First, the TE fringes on owl wings are reconfirmed to robustly suppress flow fluctuations near the TE by breaking up large TE vortices. Second, the fringes are observed to effectively suppress the shedding of wingtip vortices by mitigating the flow interaction between feathers (feather-slot interaction). These complementary mechanisms synergize to enhance the robustness and effectiveness of the TE fringe effects in owl wing models, in terms of aerodynamic force production and noise suppression. This study thus deepens our understanding of the role of TE fringes in real owl flight gliding and points to the validity and feasibility of employing owl-inspired TE fringes in practical applications of low-noise fluid machinery.


Heterogeneous fuzzy logic networks: fundamentals and development studies.
The recent trend in the development of neurofuzzy systems has profoundly emphasized the importance of synergy between the fundamentals of fuzzy sets and neural networks. The resulting frameworks of the neurofuzzy systems took advantage of an array of learning mechanisms primarily originating within the theory of neurocomputing and the use of fuzzy models (predominantly rule-based systems) being well established in the realm of fuzzy sets. Ideally, one can anticipate that neurofuzzy systems should fully exploit the linkages between these two technologies while strongly preserving their evident identities (plasticity or learning abilities to be shared by the transparency and full interpretability of the resulting neurofuzzy constructs). Interestingly, this synergy still becomes a target yet to be satisfied. This study is an attempt to address the fundamental interpretability challenge of neurofuzzy systems. Our underlying conjecture is that the transparency of any neurofuzzy system links directly with the logic fabric of the system so the logic fundamentals of the underlying architecture become of primordial relevance. Having this in mind the development of neurofuzzy models hinges on a collection of logic driven processing units named here fuzzy (logic) neurons. These are conceptually simple logic-oriented elements that come with a well-defined semantics and plasticity. Owing to their diversity, such neurons form essential building blocks of the networks. The study revisits the existing categories of logic neurons, provides with their taxonomy, helps understand their functional features and sheds light on their behavior when being treated as computational components of any neurofuzzy architecture. The two main categories of aggregative and reference neurons are deeply rooted in the fundamental operations encountered in the technology of fuzzy sets (including logic operations, linguistic modifiers, and logic reference operations). The developed heterogeneous networks come with a well-defined semantics and high interpretability (which directly translates into the rule-based representation of the networks). As the network takes advantage of various logic neurons, this imposes an immediate requirement of structural optimization, which in this study is addressed by utilizing various mechanisms of genetic optimization (genetic algorithms). We discuss the development of the networks, elaborate on the interpretation aspects and include a number of illustrative numeric examples.


Regulation of Cell Volume by Nanosecond Pulsed Electric Fields.
Stimulation of cells by nanosecond pulsed electric fields (nsPEFs) has attracted attention as a technology for medical applications such as cancer treatment. nsPEFs have been shown to affect intracellular environments without significant damage to cell membranes; however, the mechanism underlying the effect of nsPEFs on cells remains unclear. In this study, we constructed electrodes for applying nsPEFs and analyzed the change in volume of a single cell due to nsPEFs using fluorescence and Raman microscopy. It was shown that the direction of the change depended on the applied electric field; expansion due to the influx of water was observed at high electric field, and cell shrinkage was observed at low electric field. The change in cell volume was correlated to the change in the intracellular Ca2+ concentration, and nsPEFs-induced shrinking was not observed when the Ca2+-free medium was used. This result suggests that the cell shrinkage is related to the regulatory volume decrease where the cell adjusts the increase in intracellular Ca2+ concentration, inducing the efflux of ions and water from the cell.


[Effect of CD40 on Foxp3(+) Treg cell in the lung of cigarette smoke exposure mice].
To evaluate the effect of CD40 on Foxp3(+) Treg cell in the lung of cigarette smoke exposure mice. According to the random number table, 20 wild type (WT) C57 BL/6 mice and 20 CD40(-/-)C57 BL/6 mice were randomly divided into two groups: WT control group, WT smoke-exposure group (24 weeks) and CD40(-/-) control group, CD40(-/-) smoke-exposure group (24 weeks) (n=10 each). Alveolar airspace enlargement was observed by HE staining. Morphological change was evaluated by mean linear intercepts (MLI). Immunohistochemical method was used to detect the quantity of Foxp3(+) cell in the lung. The mRNA expression of Foxp3 was measured by fluorescence quantitative real-time polymerase chain reaction (qRT-PCR). The protein level of Foxp3 was measured by Western blot. Interleukin (IL)-10 and IL-35 levels in the lung were tested by enzyme-linked immunosorbent assay (ELISA). The MLI in CD40(-/-) smoke-exposure group was significantly lower than the WT smoke-exposure group[(30.0±1.7) vs (37.3±3.7) μm], but higher than the CD40(-/-) control group[(23.2±2.5) μm], WT smoke-exposure group was significantly higher than the WT control group[(22.2±1.7) μm](all P<0.05). The percentage of Foxp3(+) cell in the lungs of CD40(-/-) smoke-exposure group was significantly higher than the WT smoke-exposure group and CD40(-/-) control group[(16.89±0.75)% vs (9.65±0.74)% and (13.58±0.51)%], WT smoke-exposure group was significantly lower than WT control group[(12.13±0.81)%](all P<0.05). In the lungs, Foxp3 mRNA and protein expression in CD40(-/-) smoke-exposure group were increased compared to WT-smoke-exposure group and CD40(-/-) control group, WT smoke-exposure group were decreased compared to WT control group (all P<0.05). In the lungs, the level of IL-10 in CD40(-/-) smoke-exposure group was higher than the WT smoke-exposure group and CD40(-/-) control group[(231±25) vs (80±31) and (183±29) ng/L], WT smoke-exposure group was lower than the WT control group[(192±37) ng/L](all P<0.05). The level of IL-35 in CD40(-/-) smoke-exposure group was higher than the CD40(-/-) control group, WT control group and WT smoke-exposure group[(208±29) vs (118±29) , (148±36), (137±37) ng/L, all P<0.05]. Knockout the CD40 gene can promote the differentiation of Foxp3(+) Treg cell in the lung of cigarette smoke exposure mice, indicating that blocking the CD40-CD40 ligand pathway may contribute to alleviate the smoking-induced pulmonary emphysema.


Surgical management of transected injury to the pancreatic neck.
To present a batch of data of transected pancreatic neck injuries and to sum up the experience in surgical interventions for the injuries. We analysed 13 patients with a transected injury to the pancreatic neck from Jan. 1995 to Dec. 2000. External drainage was performed in all patients. Pancreatoduodenectomy was conducted in 2 patients with a transected injury to the pancreatic neck associated with duodenal ruptures, and TPN was administered immediately after operation. Proximal closure of the transected margin and distal pancreaticojejunostomy was performed in 4 patients. Proximal closure of the transected margin and distal pancreaticojejunostomy plus splenectomy was performed in 7 patients associated with contusion of pancreatic body or tail plus spleen rupture. 12 patients healed and one patient died of anesthetic accident during the course of restoration of the dislocation of his right hip joint. Complications occurred in 7 patients. The operation should be performed according to the degree of the injuries and associated duodenal injuries. Routine drainage and nutrient support should be recommended.


Genetics of Arthrogryposis and Macroglossia in Piemontese Cattle Breed.
Arthrogryposis and macroglossia are congenital pathologies known in several cattle breeds, including Piemontese. As variations in single genes were identified as responsible for arthrogryposis in some breeds, we decided: (i) to test the hypothesis of a similar genetic determinism for arthrogryposis in the Piemontese breed by genotyping affected and healthy animals with a high-density chip and applying genome-wide association study (GWAS), FST and canonical discriminant analysis (CDA) procedures, and (ii) to investigate with the same approach the genetic background of macroglossia, for which no genetic studies exist so far. The study included 125 animals (63 healthy, 30 with arthrogryposis, and 32 with macroglossia). Differently from what reported for other breeds, the analysis did not evidence a single strong association with the two pathologies. Rather, 23 significant markers on different chromosomes were found (7 associated to arthrogryposis, 11 to macroglossia, and 5 to both pathologies), suggesting a multifactorial genetic mechanism underlying both diseases in the Piemontese breed. In the 100-kb interval surrounding the significant SNPs, 20 and 26 genes were identified for arthrogryposis and macroglossia, respectively, with 12 genes in common to both diseases. For some genes (NTN3, KCNH1, KCNH2, and KANK3), a possible role in the pathologies can be hypothesized, being involved in processes related to muscular or nervous tissue development. The real involvement of these genes needs to be further investigated and validated.


Black art posters, an incentive to increase study enrollment among Blacks in a large cohort study.
Black art posters were offered to replace or augment the established $10 incentive for questionnaire completion in a longitudinal cohort study. Eighty-one churches located in the US southern region were divided between two intervention groups, with a control group of 24 churches from the same region. Primary outcome measures were study enrollment rates and questionnaire return rates between December 2003 and July 2004 as a proportion of church goal. 9.3% of participants returning questionnaires selected a poster in preference to $10. Half of participants offered both monetary and art incentives indicated a poster selection. Crude questionnaire return rates were 57.4% for the pooled intervention churches and 38.2% for the control churches. Enrollment rates among those offered both incentives were significantly higher (p<0.01) than when monetary incentives alone were offered after adjustment for church size, promotional dates, and average income of church members. Survey return rates were also higher in the churches offered both incentives (p=0.04). These data suggest that the black art posters improved study enrollment and survey return rates. The relatively low rate of poster selection suggests that the art primarily influenced participation indirectly, by creating a more culturally inclusive image of the study.


Endogenous Thymosin β4 Expression in Sacrococcygeal Pilonidal Sinus Disease: A Retrospective, Immunohistochemical Analysis of Excisional Skin Biopsy Samples.
Thymosin beta 4 (Tβ4) is a peptide that has been shown in dermal, corneal, and cardiac preclinical injury models to potentially affect tissue protection, regeneration, and repair. Sacrococcygeal pilonidal sinus disease (SPSD) is a chronic inflammatory disorder associated with a high incidence of recurrence, chronic fistulation, and a challenging postoperative surgical wound healing process. Retrospectively, an immunohistochemical analysis was conducted to evaluate endogenous Tβ4 expression in excisional skin biopsies from patients with SPSD. Patient demographics (age, gender) and surgical procedure data were obtained from their electronic medical records. Two (2) samples were cut from each specimen and prepared for histopathological assessment: 1 from the inflamed sinus tracts containing hair and granulation tissue (chronic wound group) and 1 from the normal tissue at least 1 cm away from the sinus tract (control group). Tβ4 expression was evaluated in the epidermal, dermal/subcutaneous collagen, and vascular structures of the samples from the sinus tract and healthy tissue. Inflamed sinus tract tissue and noninflamed normal tissue adjacent to the sinus tract were sampled from each specimen to confirm the diagnosis of SPSD and to determine distribution and intensity of Tβ4 expression. Presence of cytoplasmic staining for Tβ4 was considered in favor of positive Tβ4 expression; intensity of Tβ4 expression was scored as 0 = no staining, 1 = mild, 2 = moderate, and 3 = strong level of expression. A total of 31 excisional skin biopsy specimens were available from 31 patients with SPSD (mean age 26.0 ± 7.6 years, 25 [80.6%] men, 6 [19.4%] women) who underwent primary surgical closure. Demographic variables were analyzed using descriptive statistics. Data compliance with normal distribution was evaluated using the Kolmogorov-Smirnov and Shapiro-Wilk tests, and the Mann-Whitney U test was used for comparison of numerical data. P <.05 was considered statistically significant. Inflamed sinus tract tissue had significantly higher Tβ4 expression scores than noninflamed tissue samples in the epidermis (2.4 ± 0.8 [1.0-3.0] versus 0.8 ± 0.5 [0.0-2.0], P = .000), dermal/subcutaneous collagen (2.6 ± 0.5 [2.0-3.0] versus 1.6 ± 0.5 [1.0-2.0], P = .000), and vascular structure (2.6 ± 0.5 [2.0-3.0] versus 1.1 ± 0.3 [1.0-2.0], P = .000). Study findings indicate Tβ4 is endogenously expressed in normal skin tissue and is overexpressed in inflamed sinus tract tissue in patients with SPSD. Preclinical studies with a larger sample size are needed to enhance understanding of the potential role of Tβ4 in the inflammatory and tissue remodeling processes of SPSD by elucidating its mechanism of action at the molecular level, physiological role, and the therapeutic potential in dermal healing.


Benthic impact assessment of a dredge sediment disposal in a dynamic resilient environment.
The eastern part of the Bay of Seine (English Channel) is highly impacted by harbour activities and the dumping of dredged sediment by the port authorities of Le Havre (GPMH) and Rouen (GPMR). Sediment dredged by the GPMH (2-2.5 millions of m3 per year) has been disposed at the subtidal Octeville site since 1947. Since the 2000s, mainly fine sediment (80% of fine particles <63 μm) has been disposed using alternate mosaic boxes with limited thickness (0.2 to 0.6 m per box per year), preventing the accumulation of disposed sediment in some parts of the dumping site. During the period August 2016 to September 2017, an experimental study was set up to identify the spatio-temporal changes of the macrobenthos collected at ten stations on six dates: three stations where different volumes of sediments were dumped (from 41,000 to 186,000 m3), two stations located within the Octeville site but without dumping operations during the study and five stations outside the disposal site (northern and southern zones). The Taxonomic Richness, total abundance and abundances of the polychaete Owenia fusiformis show negative correlations with the volumes of deposited sediment, whereas use of the AMBI (AZTI's Marine Biotic Index) and B2OA (Benthic Opportunist Annelids Amphipods) indices does not allow us to distinguish the impacted zone. Our study shows that the impact of dumping remains local and the benthic habitats display a high degree of resilience with rapid recovery of the community after the cessation of disturbance. Recommendations on the future long-term strategy are proposed to improve assessment and minimise the impact of dumping sediment on this benthic habitat.


Habitat partitioning during character displacement between the sexes.
Ecological differences between the sexes are often interpreted as evidence of within-species ecological character displacement (ECD), a hypothesis with almost no direct tests. Here, we experimentally test two predictions that are direct corollaries of ECD between the sexes, in a salamander. First, we find support for the prediction that each sex has a growth rate advantage in the aquatic microhabitat where it is most commonly found. Second, we test the prediction that selection for ECD in the breeding environment may affect partial migration out of this environment. We found that phenotype-dependent migration resulted in a shift in the phenotypic distribution across treatments, with the highest sexual dimorphism occurring among residents at high founding density, suggesting that migration and ECD can both be driven by competition. Our work illustrates how complex patterns of habitat partitioning evolve during ECD between the sexes and suggest ECD and partial migration can interact to effect both ecological dynamics and evolution of sexual dimorphism.


Developmentally regulated RNA transcripts coding for alcohol dehydrogenase in Drosophila affinidisjuncta.
The organization of the gene coding for alcohol dehydrogenase (Adh) in Drosophila affinidisjuncta has been determined by physically mapping Adh RNA transcripts to cloned genomic DNA. Two distinct transcript types accumulate with developmental specificity. Because only a single genomic Adh locus is detected in D. affinidisjuncta, and since all Adh transcripts appear to be identical except at their termini, the two Adh RNA types are products of the same gene. One type of transcript, abundant in adults, contains a small 5' terminal exon that is completely lacking in the other type of transcript, which accumulates in larvae. This 5' end difference suggests that the D. affinidisjuncta Adh gene, like the homologous gene from the distantly related species D. melanogaster, is expressed from two promoters. According to the transcription map, these D. affinidisjuncta promoters are separated by approximately 560 base pairs of genomic DNA sequence. D. affinidisjuncta Adh transcripts also resemble D. melanogaster Adh transcripts in both their overall organization and their developmental distribution. Multiple 3' ends are responsible for the size heterogeneity of both types of D. affinidisjuncta Adh RNA, and some of these also appear with stage specificity.


Elevated olivine weathering rates and sulfate formation at cryogenic temperatures on Mars.
Large Hesperian-aged (~3.7 Ga) layered deposits of sulfate-rich sediments in the equatorial regions of Mars have been suggested to be evidence for ephemeral playa environments. But early Mars may not have been warm enough to support conditions similar to what occurs in arid environments on Earth. Instead cold, icy environments may have been widespread. Under cryogenic conditions sulfate formation might be blocked, since kinetics of silicate weathering are typically strongly retarded at temperatures well below 0 °C. But cryo-concentration of acidic solutions may counteract the slow kinetics. Here we show that cryo-concentrated acidic brines rapidly chemically weather olivine minerals and form sulfate minerals at temperatures as low as -60 °C. These experimental results demonstrate the viability of sulfate formation under current Martian conditions, even in the polar regions. An ice-hosted sedimentation and weathering model may provide a compelling description of the origin of large Hesperian-aged layered sulfate deposits on Mars.


Management of cancer during pregnancy.
Although cancer during pregnancy is infrequent, its management is difficult for patients, their families, and their physicians. When termination of the pregnancy is unacceptable, decisions regarding the use of irradiation and chemotherapy are complicated by the well-known high risks of abortion and fetal malformation. This risk is concentrated in the first trimester and varies with the choice of chemotherapeutic agents or combinations of agents. There is only minimal evidence of increased risk of malformation or abortion in the second or third trimester. Recent progress in cancer therapy has made cure a reasonable goal, and for some malignant neoplasms, cure is still possible even when initial therapy is modified or delayed. When cure is a reasonable goal, curative therapy should not be compromised by modification or delay. When treatment for cure or significant palliation is not possible, however, the goal should shift to protection of the fetus from damage by the injudicious use of teratogenic cancer therapy. This report will review the available data that may assist in these difficult decisions.


On the algorithm for computing body surface Laplacians in an inhomogeneous volume conductor of arbitrary shape.
In this short communication, a corrected version of algorithm for calculating the body surface Laplacian (BSL) in an inhomogeneous volume conductor of arbitrary shape is described. The present computer simulation results show that the corrected algorithm can give an accurate numerical solution. This algorithm was then applied to examine the effect of the lung inhomogeneity on the BSL's in a realistically shaped torso model. The simulation results show that the low-conductivity lungs have little effect on both topology and magnitudes of body surface Laplacian maps (BSLM's) for a single dipole source.


Mediterranean Alcohol-Drinking Patterns and All-Cause Mortality in Women More Than 55 Years Old and Men More Than 50 Years Old in the "Seguimiento Universidad de Navarra" (SUN) Cohort.
Most of the available epidemiological evidence on alcohol and chronic disease agrees on recommending alcohol abstention to young people, but some controversy exists about the most appropriate recommendation for alcohol abstention for people of older ages. A growing body of evidence suggests that the pattern of alcohol consumption is likely to be a strong effect modifier. The Mediterranean Alcohol Drinking Pattern (MADP) represents a score integrating several dimensions of drinking patterns (moderation, preference for red wine, drinking with meals, and avoiding binge drinking). Our aim was to clarify this issue and provide more precise recommendations on alcohol consumption. We prospectively followed-up 2226 participants (men older than 50 years and women older than 55 years at baseline) in the Seguimiento Universidad de Navarra (SUN) cohort. We classified participants into three categories of adherence to the MADP score (low, moderate, and high), and we added a fourth category for abstainers. Cox regression models estimated multivariable-adjusted hazard ratios (HR) of all-cause death and 95% confidence intervals (CI) using low MADP adherence as the reference category. The strongest reduction in risk of mortality was observed for those with high adherence to the MADP, with an HR of 0.54 (95% CI: 0.37-0.80). The moderate adherence group (HR = 0.65, 95% CI: 0.44-0.96) and the abstention group (HR = 0.60, 95% CI: 0.36-0.98) also exhibited lower risks of mortality than the low MADP adherence group. based on the available evidence, a public health message can be provided to people older than 50 years as follows: among those who drink alcohol, high adherence to the MADP score could substantially reduce their risk of all-cause mortality.


Primary definitive nasal correction in patients presenting for late unilateral cleft lip repair.
Almost 25 percent of unilateral cleft lip and palate patients present with their deformity in their teens or later years in the developing world. Because more than 80 percent of the world population lives in the developing world, the established protocol for repair of these deformities is not applicable to these patients. Despite the magnitude, there are no significant reports in the literature that deal with this problem. Several issues need to be addressed, but the author limits himself here to the correction of the nasal deformity. The patients at this age are very much concerned with the aesthetic outcome. Procedures described hitherto for primary nasal correction in infants are not successful in restoring nasal shape and symmetry at this late age of presentation. Our experience with radical correction of secondary nasal deformity in unilateral cleft lip patients presenting late prompted us to extend the concept by undertaking a definitive primary correction of the nasal deformity in cleft patients presenting late. Twenty-two patients with unilateral cleft lip deformity (nine male patients and 13 female patients) with ages ranging from 13 to 22 years, presenting between August of 1997 and December of 2000, are included in this study. Of these, 11 patients had a cleft of the lip alone, eight also had a cleft of the alveolus, and three had a cleft of the palate continuous with the cleft lip. All patients showed some maxillary hypoplasia. An external rhinoplasty with lip repair was carried out in all patients. The corrective procedures on the nose included columellar lengthening; augmentation along the pyriform margin, nasal floor, and alveolus using bone grafts; submucous resection of the nasal septum; repositioning of lower lateral cartilages; and augmentation of nasal dorsum by bone graft. Clinical follow-up ranged from 4 to 24 months, and the median follow-up period was 13 months. Results have been very good, and much better than results seen earlier with other primary rhinoplasty techniques. While repairing unilateral cleft lip in adolescents, the author thinks it would be most appropriate to address the entire gamut of the deformity in a single stage, provide complete vector reorientation, and augment the hypoplastic elements by autologous tissue. It is not just the fear of poor follow-up, but that merely correcting the lip deformity in these patients without attempting definitive rhinoplasty, in the author's opinion, would be insufficient surgical intervention.


Correction of half the cardiomyocytes fully rescue Friedreich ataxia mitochondrial cardiomyopathy through cell-autonomous mechanisms.
Friedreich ataxia (FA) is currently an incurable inherited mitochondrial neurodegenerative disease caused by reduced levels of frataxin. Cardiac failure constitutes the main cause of premature death in FA. While adeno-associated virus-mediated cardiac gene therapy was shown to fully reverse the cardiac and mitochondrial phenotype in mouse models, this was achieved at high dose of vector resulting in the transduction of almost all cardiomyocytes, a dose and biodistribution that is unlikely to be replicated in clinic. The purpose of this study was to define the minimum vector biodistribution corresponding to the therapeutic threshold, at different stages of the disease progression. Correlative analysis of vector cardiac biodistribution, survival, cardiac function and biochemical hallmarks of the disease revealed that full rescue of the cardiac function was achieved when only half of the cardiomyocytes were transduced. In addition, meaningful therapeutic effect was achieved with as little as 30% transduction coverage. This therapeutic effect was mediated through cell-autonomous mechanisms for mitochondria homeostasis, although a significant increase in survival of uncorrected neighboring cells was observed. Overall, this study identifies the biodistribution thresholds and the underlying mechanisms conditioning the success of cardiac gene therapy in Friedreich ataxia and provides guidelines for the development of the clinical administration paradigm.


Elaboration of telomeres in yeast: recognition and modification of termini from Oxytricha macronuclear DNA.
The termini of macronuclear DNA molecules from the protozoan Oxytricha fallax share a common sequence and structure, both of which differ markedly from those deduced for yeast telomeres. Despite these differences, terminal restriction fragments from O. fallax macronuclear DNA can support telomere formation in yeasts. Two linear plasmids (LYX-1 and LYX-2) constructed by ligating BamHI-digested total Oxytricha macronuclear DNA to a yeast vector were analyzed. One end of LYX-1 and both ends of LYX-2 are derived from the Oxytricha DNA that encodes rRNA (rDNA) whereas the other end of LYX-1 is from an Oxytricha fragment other than rDNA. After propagation in yeast, both ends of LYX-1 and LYX-2 retain the C4A4 repeat characteristic of the O. fallax terminal sequence. In addition, both ends of both plasmids acquire 300-1000 base pairs of DNA containing the sequence (C-A)n, a sequence found near the termini of yeast chromosomes. Thus, at least two different Oxytricha termini display distinctive properties in yeast cells in that linear plasmids containing them are not degraded nor are they integrated into chromosomal DNA. These Oxytricha termini may act directly as telomeres in yeast; alternatively, the Oxytricha DNA may serve as a signal that results in the elaboration of a yeast telomere on the ciliate DNA.


Molecular genetics of peripheral neuropathies.
Molecular genetic techniques have identified the mutational basis of many inherited neuropathies due to metabolic disorders, including porphyrias, leukodystrophies and other storage diseases. Inherited amyloid neuropathies may be caused by mutations in three different genes: those for gelsolin, apolipoprotein A1 and, most frequently, transthyretin. The classification of hereditary motor and sensory neuropathies has been changed by the identification of underlying mutations encoding myelin proteins, P0 and peripheral myelin protein 22 and connexin 32. Peripheral myelin protein 22 gene defects are also seen in hereditary liability to pressure palsies. There is peripheral sensory nerve involvement in one of the diseases caused by an unstable trinucleotide repeat sequence: X-linked bulbospinal neuronopathy. These advances can be translated into clinical practice, leading to improved diagnosis and genetic counselling.


Labeling Laws for Personal Care Products: Potential Pitfalls for The Consumer.
Consumers of personal care products must be made aware of the potential pitfalls that arise from current labeling practices and regulations. For example, terms such as "hypoallergenic", "sensitive skin", and "baby product" lack legal definitions, and terms such as "fragrance-free" are frequently misinterpreted. Personal care products are a general category that includes such items as cosmetics, drugs, dietary supplements, and consumer goods. This overview of current US FDA regulations of products in this category reviews pertinent regulatory policies and highlights potential consumer pitfalls when evaluating product labels. In particular, current labeling laws permit the use of language that may be incomplete, misinterpreted, or applied solely for marketing purposes. It is important that consumers understand the meaning of labeling terms, the regulations that govern them, and especially understand the potential pitfalls related to these terms.


What is needed for informed decisions about prostate cancer screening: perspectives of African-American and Hispanic men.
Professional guidelines suggest that men should learn about risks and benefits of screening to make informed decisions consistent with their preferences. We used concept mapping to investigate views of informed decision making (IDM) of minority men. Statements about what men need for IDM about prostate cancer screening were sorted by similarity and rated for importance by 16 Hispanic and 15 African-American men. Multidimensional scaling and cluster analysis were used to develop a concept map for IDM. The 10-cluster solution was selected. The clusters rated most important were labeled Future Considerations, What to Know and Decision to Make. Clusters labeled Social Support and Sharing Perspectives depicted social aspects of the decision and were intermediate in importance. There was strong correlation in relative importance ratings of clusters by African-American and Hispanic men. However, African-American men gave higher importance ratings than Hispanic men. Concept mapping, a method with strong participatory elements, was useful in identifying conceptual frameworks for IDM of African-American and Hispanic men. Health education to support IDM requires some shifts in focus and strategy. It is important that interventions with minority men build upon a strong conceptual framework.


Insights from letter position dyslexia on morphological decomposition in reading.
We explored morphological decomposition in reading, the locus in the reading process in which it takes place and its nature, comparing different types of morphemes. We assessed these questions through the analysis of letter position errors in readers with letter position dyslexia (LPD). LPD is a selective impairment to letter position encoding in the early stage of word reading, which results in letter migrations (such as reading "cloud" for "could"). We used the fact that migrations in LPD occur mainly in word-interior letters, whereas exterior letters rarely migrate. The rationale was that if morphological decomposition occurs prior to letter position encoding and strips off affixes, word-interior letters adjacent to an affix (e.g., signs-signs) would become exterior following affix-stripping and hence exhibit fewer migrations. We tested 11 Hebrew readers with developmental LPD and 1 with acquired LPD in 6 experiments of reading aloud, lexical decision, and comprehension, at the single word and sentence levels (compared with 25 age-matched control participants). The LPD participants read a total of 12,496 migratable words. We examined migrations next to inflectional, derivational, or bound function morphemes compared with migrations of exterior letters. The results were that root letters adjacent to inflectional and derivational morphemes were treated like middle letters, and migrated frequently, whereas root letters adjacent to bound function morphemes patterned with exterior letters, and almost never migrated. Given that LPD is a pre-lexical deficit, these results indicate that morphological decomposition takes place in an early, pre-lexical stage. The finding that morphologically complex nonwords showed the same patterns indicates that this decomposition is structurally, rather than lexically, driven. We suggest that letter position encoding takes place before morphological analysis, but in some cases, as with bound function morphemes, the complex word is re-analyzed as two separate words. In this reanalysis, letter positions in each constituent word are encoded separately, and hence the exterior letters of the root are treated as exterior and do not migrate.


Induction of oxidative DNA damage by Helicobacter pylori in HT29 cells.
Infection with Helicobacter pylori has been associated with the development of gastric adenocarcinoma in humans. Several routes have been implicated, the main one being oxidative DNA damage resulting from chronic inflammation, which accompanies infection. However, DNA has been demonstrated in human cells after in vitro incubation with H. pylori sonicates. Using the fragment length analysis using restriction enzymes (FLARE) assay, this study investigates the DNA damaging potential of three clinical isolates of H. pylori on cultured HT29 cells. Significant amounts of oxidative DNA damage were detected in HT29 cells following a 72-hour incubation with each H. pylori isolate. As tumour induction is a known consequence of oxidative DNA damage, chronic infection with the organism may lead to the development of adenocarcinoma of the stomach.


Schizophrenia-Associated MIR204 Regulates Noncoding RNAs and Affects Neurotransmitter and Ion Channel Gene Sets.
As regulators of gene expression, microRNAs (miRNAs) are likely to play an important role in the development of disease. In this study we present a large-scale strategy to identify miRNAs with a role in the regulation of neuronal processes. Thereby we found variant rs7861254 located near the MIR204 gene to be significantly associated with schizophrenia. This variant resulted in reduced expression of miR-204 in neuronal-like SH-SY5Y cells. Analysis of the consequences of the altered miR-204 expression on the transcriptome of these cells uncovered a new mode of action for miR-204, being the regulation of noncoding RNAs (ncRNAs), including several miRNAs, such as MIR296. Furthermore, pathway analysis showed downstream effects of miR-204 on neurotransmitter and ion channel related gene sets, potentially mediated by miRNAs regulated through miR-204.


Structure, Magnetism, and the Interaction of Water with Ti-Doped Fe3O4 Surfaces.
The functionality of magnetite, Fe3O4, for catalysis and spintronics applications is dependent on the molar ratio of Fe2+ and Fe3+ and their distribution at the surface. In turn, this depends on a poorly understood interplay between crystallographic orientation, dopants, and the reactive adsorption of atmospheric species such as water. Here, (100)-, (110)-, and (111)-oriented films of titano-magnetite, Fe(3-x)TixO4, were grown by pulsed laser deposition and their composition, valence distribution, magnetism, and interaction with water were studied by ambient pressure X-ray photoelectron spectroscopy (AP-XPS) and X-ray magnetic circular dichroism. Although the bulk compositions match the desired stoichiometry, the surfaces were found to be enriched in Ti4+, especially the top 1 nm. The highest surface energy (110) film was the most reduced, tied to local Ti enrichment, and a corresponding decreased magnetic moment. AP-XPS showed that incorporation of x = 0.25 Ti dramatically lowered the propensity to form hydroxyl species at a given relative humidity, and also that hydroxylation is relatively invariant with orientation. In contrast, the affinity for water is similar across orientations, regardless of Ti incorporation, suggesting that relative humidity controls its uptake. The findings may help demystify the interactions that lead to specific distributions of Fe2+ and Fe3+ at magnetite surfaces, toward design of more deliberately active catalysts and magnetic devices.


Differentiating therapy in acute myeloid leukemia.
Differentiating therapy is a new antineoplastic strategy which has received increasing attention due to the remarkable activity of the vitamin A derivative, all-trans retinoic acid (ATRA) in patients with acute promyelocytic leukemia (APL). Although it has been known for years that a variety of agents, including retinoids, could induce leukemic cells to differentiate in vitro, it was not until the initial report from Shanghai in 1988 that laboratory studies translated into clinical activity and benefit in patients. Since this initial report, a number of studies have confirmed that the majority of patients with both newly diagnosed and previously chemotherapy-treated patients with APL achieve complete remission (CR) with ATRA. In addition, the characteristic life-threatening coagulopathy resolves quickly. Several limitations to this approach have emerged, including the development of retinoid resistance, hyperleukocytosis and the retinoic acid syndrome, a constellation of findings including unexplained fever, fluid retention, pleuropericardial effusions and pulmonary infiltrates. Although ATRA is very effective in inducing CR, its benefits compared to conventional chemotherapy are only now being addressed. The first prospective randomized trial comparing ATRA plus chemotherapy to chemotherapy alone was terminated early because of an improved event-free survival for patients receiving ATRA. The benefit was attributable to a difference in relapse rate. A large, intergroup, prospective, randomized trial comparing conventional chemotherapy to ATRA for induction and ATRA to observation for maintenance has recently completed accrual and will provide insight into the emerging role of ATRA in patients with APL. ATRA represents the first example of a specific form of antileukemic therapy targeting a specific genetic abnormality and may serve as a paradigm for the development of differentiating therapy for patients with other hematologic malignancies.


[An objective index of quality and prognostication of the storage time for cod liver oil].
Through a series of tests an objective indicator of the degree of the cod oil oxidation-the content of aldehydes reacting with benzidine-has been established. Mathematic processing of experimental data helped determine the limits of the aldehydes content in the oil of different quality, while the maximum permissible standards were fixed following biological tests carried out by the Institute of Nutrition of the AMS of the USSR. These data and investigations into the quality of the oil during its preparation and subsequent storage under different conditions formed a background to be establishing maximum permissible content of aldehydes in the half-finished and final products. Through experiments there were found the increment of aldehydes and the fall of the A-vitamin activity over a storage period of one month in large bottles of 9 1 capacity at temperatures of 10+/-2 and 20+/-2 degrees. The available data enabled it to deduce a formula for fixing approximate storage periods for medicinal oil both on shipment from the production site and at the time of control checking of its quality. The maximum permissible content of aldehydes in the medicinal oil and the method of establishing storage time are recommended to be included into the corresponding intem of the state standards-GOST, or in the pharmacopeia.


Minimalistic predictor of protein binding energy: contribution of solvation factor to protein binding.
It has long been known that solvation plays an important role in protein-protein interactions. Here, we use a minimalistic solvation-based model for predicting protein binding energy to estimate quantitatively the contribution of the solvation factor in protein binding. The factor is described by a simple linear combination of buried surface areas according to amino-acid types. Even without structural optimization, our minimalistic model demonstrates a predictive power comparable to more complex methods, making the proposed approach the basis for high throughput applications. Application of the model to a proteomic database shows that receptor-substrate complexes involved in signaling have lower affinities than enzyme-inhibitor and antibody-antigen complexes, and they differ by chemical compositions on interfaces. Also, we found that protein complexes with components that come from the same genes generally have lower affinities than complexes formed by proteins from different genes, but in this case the difference originates from different interface areas. The model was implemented in the software PYTHON, and the source code can be found on the Shakhnovich group webpage: http://faculty.chemistry.harvard.edu/shakhnovich/software.


Using palliative care assessment tools to influence and enhance clinical practice.
The Palliative Care Outcomes Collaboration focuses on supporting palliative care services in Australia to measure service- and patient-level outcomes, and to use these data to drive continuous quality improvement. The introduction of a suite of clinical assessment tools nationally has resulted in important enhancements to care provision at individual service level. Improved communication, enhanced assessment of patient needs, and improved identification of triggers for the need to change care plans or for referral have resulted from this change.


Ecological dynamics and large scale phenotypic differentiation in density-dependent populations.
Spatial differentiation of phenotypes is assumed to be determined by a combination of fluctuating selection producing adaptations to the local environment and a homogenizing effect of migration. We present a model with density regulation and a density-dependent fitness function affected by spatio-temporal variability in population size driven by spatially correlated fluctuations in the environment causing fluctuating r- and K-selection on a set of traits. We derive the variance in local mean phenotypes and show how the spatial scales of the correlations between the components of the mean phenotype depend on ecological parameters. The degree of spatial differentiation of phenotypes is strongly influenced by parameters affecting ecological dynamics. In the case of a one-dimensional character the geographical scale of variation in the mean phenotype has simply an additive term corresponding to the Moran effect in population dynamics as well as a term determined by dispersal and strength of local selection. The degree of phenotypic differentiation increases with decreasing strength of local density dependence and decreasing strength of local selection. These results imply that the form of the spatial autocorrelation function can reveal important information about ecological and evolutionary processes causing phenotypic differentiation in space.


[Diagnostic capabilities of single-photon emission computed tomography/computed tomography in the evaluation of perfusion disorders in pulmonary hypertension].
to study the diagnostic capabilities of single-photon emission computed tomography/computed tomography (SPECT/ CT) in the evaluation of lung perfusion in pulmonary hypertension (PH). Ten patients with pulmonary hypertension were examined. Its diagnosis was verified in accordance with the Russian guidelines for the diagnosis and treatment of PH, by conducting a set of studies involving right heart catheterization. CT angiopulmonography was carried out in all the patients. At the same time all the patients underwent a radiological study on a combined SPECT/CT scanner with the intravenous injection of the radiotracer 9mTc-MAA (technetium-99m-labelled macroaggregated albumin) with an activity of 2-4 mCi. CT-angiopulmonography revealed no thrombi in the pulmonary artery lumen in 7 patients and confirmed the diagnosis of pulmonary thromboembolism (PTE) at the level of segmental vessels in 3. SPECT/CT examination identified 5 cases of non-thromboembolic PH and 5 cases of PTE with consideration for SPECT perfusion defects and no visible lung tissue (fibrous or other) changes on a CT scan. It should be noted that the 5 patients diagnosed as having PTE included 2 patients in whom PTE had not previously been diagnosed at CT angiopulmonography. This may be due to that this technique cannot always clearly visualize thrombi in the pulmonary vessel lumen at a subsegmental level. The inivestigations have indicated that, by clearly ivisualizing pulmonary aanatomy, morphology, and function, combined SPECT/CT imaging is a valuable diagnostic tool in detecting lung pathology.


Membrane-Associated α-Tubulin Is Less Acetylated in Postmortem Prefrontal Cortex from Depressed Subjects Relative to Controls: Cytoskeletal Dynamics, HDAC6, and Depression.
Cytoskeletal proteins and post-translational modifications play a role in mood disorders. Post-translational modifications of tubulin also alter microtubule dynamics. Furthermore, tubulin interacts closely with Gαs, the G-protein responsible for activation of adenylyl cyclase. Postmortem tissue derived from depressed suicide brain showed increased Gαs in lipid-raft domains compared with normal subjects. Gαs, when ensconced in lipid rafts, couples less effectively with adenylyl cyclase to produce cAMP, and this is reversed by antidepressant treatment. A recent in vitro study demonstrated that tubulin anchors Gαs to lipid rafts and that increased tubulin acetylation (due to HDAC6 inhibition) and antidepressant treatment decreased the proportion of Gαs complexed with tubulin. This suggested that deacetylated-tubulin might be more prevalent in depression. This study examined tubulin acetylation in whole-tissue homogenate, plasma membrane, and lipid-raft membrane domains in tissue from normal control subjects, depressed suicides, and depressed nonsuicides (human males/females). While tissue homogenate showed no changes in tubulin acetylation between control, depressed suicides, and depressed nonsuicides, plasma membrane-associated tubulin showed significant decreases in acetylation from depressed suicides and depressed nonsuicides compared with controls. No change was seen in expression of the enzymes responsible for tubulin acetylation or deacetylation. These data suggest that, during depression, membrane-localized tubulin maintains a lower acetylation state, permitting increased sequestration of Gαs in lipid-raft domains, where it is less likely to couple to adenylyl cyclase for cAMP production. Thus, membrane tubulin may play a role in mood disorders, which could be exploited for diagnosis and treatment.SIGNIFICANCE STATEMENT There is little understanding about the molecular mechanisms involved in the development of depression and, in severe cases, suicide. Evidence for the role of microtubule modifications in progression of depressive disorders is emerging. These postmortem data provide strong evidence for membrane tubulin modification leading to reduced efficacy of the G protein, Gαs, in depression. This study reveals a direct link between decreased tubulin acetylation in human depression and the increased localization of Gαs in lipid-raft domains responsible for attenuated cAMP signaling. The evidence presented here suggest a novel diagnostic and therapeutic locus for depression.


Crystallization of cytochromes P450 and substrate-enzyme interactions.
Our understanding of structure-function relationships have made considerable advances owing to the increasing number of new P450 crystal structures. This is especially true with mammalian P450s. As always, the main bottleneck in a structure determination project is crystallization. While the crystallization techniques used for P450 crystal growth are not much different from that utilized for other proteins, special protein engineering strategies have been developed in order to generate soluble, homogeneous membrane-bound P450 samples amendable for crystallization. Newly determined P450 structures also provide convincing evidence that P450 enzymes are highly dynamic and flexible. Common structural elements found in all P450s have been identified that undergo large conformational changes to allow substrate access and product release. In addition, flexible regions may enable the active site to adapt to the binding of substrates of different size, shape, and polarity. This review will focus on the successful membrane P450 crystallization techniques and the new structural insights based on the growing P450 structure database.


A modular cloning system for standardized assembly of multigene constructs.
The field of synthetic biology promises to revolutionize biotechnology through the design of organisms with novel phenotypes useful for medicine, agriculture and industry. However, a limiting factor is the ability of current methods to assemble complex DNA molecules encoding multiple genetic elements in various predefined arrangements. We present here a hierarchical modular cloning system that allows the creation at will and with high efficiency of any eukaryotic multigene construct, starting from libraries of defined and validated basic modules containing regulatory and coding sequences. This system is based on the ability of type IIS restriction enzymes to assemble multiple DNA fragments in a defined linear order. We constructed a 33 kb DNA molecule containing 11 transcription units made from 44 individual basic modules in only three successive cloning steps. This modular cloning (MoClo) system can be readily automated and will be extremely useful for applications such as gene stacking and metabolic engineering.


The association between olfactory impairment and total mortality in older adults.
Population-based data on the relationship between impaired olfaction and risk of mortality among older adults are lacking. We used a representative cohort of adults aged 60 years or older to assess whether olfactory loss is a predictor of mortality, independent of potential confounders. Olfaction was measured by the San Diego Odor Identification Test (SDOIT) among 1,636 participants enrolled in the Blue Mountains Eye Study (2002-2004). Five-year all-cause mortality was confirmed using the Australian National Death Index. More than one in five participants (21.8%) with olfactory impairment had died over the 5 years compared with less than 10% of participants without olfactory loss. Moderate olfactory loss (SDOIT score ≤3) was associated with a 68% increased risk of all-cause mortality (multivariable-adjusted hazard ratio, 1.68; 95% confidence interval, 1.10-2.56). This association did not persist after further adjustment for cognitive impairment. The association between olfactory loss and all-cause mortality was more marked among older participants (≥70 years) than younger participants (<70 years) with olfactory impairment (multivariable-adjusted hazard ratio, 1.48; 95% confidence interval, 1.02-2.15). However, adjusting for cognitive impairment diminished this association. Older adults with moderately impaired olfaction compared with those with normal olfaction had a higher risk of dying 5 years later. The relationship between olfaction and mortality, however, may be largely mediated by cognitive impairment in these older adults. Our findings highlight the value of identifying olfactory loss in the preclinical stage in the older patient before the development of related comorbidities.


Anti-asthmatic effects of Perilla seed oil in the guinea pig in vitro and in vivo.
The aim of this study was to investigate the anti-asthmatic effects of Perilla seed oil in vitro and in vivo in sensitized guinea pigs. Aerosolized antigen caused an immediate bronchoconstriction. Perilla seed oil per os inhibited the increase in lung resistance and the decrease in dynamic lung compliance in a dose-dependent manner with an ED50 (95 % confidence interval, CI) of 1.10 (0.98 - 1.24) g/kg and 1.07 (0.94 - 1.22) g/kg, respectively. Infiltration of leukocytes, mononuclear cells, eosinophils and neutrophils induced by inhaling antigen was also inhibited by Perilla seed oil in a dose-dependent manner with an ED50 (95 % CI) of 1.00 (0.86 - 1.15), 1.24 (1.10 - 1.38), 0.63 (0.51 - 0.77) and 0.61 (0.38 - 0.98) g/kg, respectively. Perilla seed oil (5 - 500 microg/mL) inhibited the slow reaction substance of anaphylaxis (SRS-A) release induced by antigen challenge in lung tissue of sensitized guinea pigs. It also inhibited calcium ionophore (A(23187))-induced leukotriene (LT) D4 release from the lung tissue of non-sensitized guinea pigs in a concentration-dependent manner with an IC50 (95 % CI) of 50 (36 - 69) microg/mL. These results indicate that Perilla seed oil may improve lung function in asthma by controlling eicosanoid production and suppressing LT generation.


Contingent Work Rising: Implications for the Timing of Marriage in Japan.
Employment has become increasingly precarious in developed countries, meaning that, for many young adults, jobs provide neither benefits nor security, more work is part time, and employers are increasingly hiring workers from temporary help agencies and contract companies rather than as employees of their own company. These changes in employment relations have profound effects on gender roles and on family transitions of young adults, especially young men and in particular in countries such as Japan, where there are rigid family norms and the male-breadwinner tradition still prevails. The authors examined the effects of the experience of non-regular work on the timing of marriage and whether this differs by sex. Using recent life history data from Japan, they found that men working in non-regular jobs are especially likely to postpone marriage. The implications of the growth of precarious work for changes in work and family institutions in Japan are discussed.


Lifting the lid on geographic complexity in the relationship between body mass index and education in China.
In China, rising obesity has coincided with increasing affluence. Few studies have properly accounted for geographic variation, however, which may influence prior results. Using large data with biomarkers in China, we show body mass index (BMI) to be positively correlated with higher person-level education if estimated using ordinary least squares. In stark contrast, fitting the same data within a multilevel model gives the complete opposite result. We go on to show that the relationship between BMI and person-level education in China is dependent upon geography, underlining why multilevel modelling is crucial for revealing these types of people-place contingencies.


Hippocampus-related cognitive impairments in young apoE4 targeted replacement mice.
We presently investigated the effects of apolipoprotein E4 (apoE4), the most prevalent genetic risk factor for Alzheimer's disease, on the cognitive performance of young targeted replacement apoE4 mice. We revealed that these mice were impaired in the object recognition and Morris water maze tests, both of which are associated with hippocampal learning and memory, relative to that of the apoE3 mice. These results are consistent with previous histological and biochemical findings that hippocampal neurons are specifically affected by apoE4. The suggestion that the behavioral impairments of the apoE4 mice are related to the hippocampal neuropathology of these mice is further supported by the fear conditioning test. This test revealed that the performance of the apoE4 mice in the contextual component, which is hippocampus related, was impaired, whereas their cued test response, which is amygdala driven, was not. The stress levels of the apoE4 and apoE3 mice, as unraveled by the light/dark anxiety test, were similar, suggesting that the observed cognitive impairments of the apoE4 mice are not related to differences in the basal anxiety levels of these mice. In conclusion, the present study shows that young apoE4 targeted replacement mice are impaired in numerous hippocampus-related learning and memory tasks.


BRCA2 controls DNA:RNA hybrid level at DSBs by mediating RNase H2 recruitment.
DNA double-strand breaks (DSBs) are toxic DNA lesions, which, if not properly repaired, may lead to genomic instability, cell death and senescence. Damage-induced long non-coding RNAs (dilncRNAs) are transcribed from broken DNA ends and contribute to DNA damage response (DDR) signaling. Here we show that dilncRNAs play a role in DSB repair by homologous recombination (HR) by contributing to the recruitment of the HR proteins BRCA1, BRCA2, and RAD51, without affecting DNA-end resection. In S/G2-phase cells, dilncRNAs pair to the resected DNA ends and form DNA:RNA hybrids, which are recognized by BRCA1. We also show that BRCA2 directly interacts with RNase H2, mediates its localization to DSBs in the S/G2 cell-cycle phase, and controls DNA:RNA hybrid levels at DSBs. These results demonstrate that regulated DNA:RNA hybrid levels at DSBs contribute to HR-mediated repair.


Changes in tissue blood flow and beta-receptor density of skeletal muscle in rats treated with the beta2-adrenoceptor agonist clenbuterol.
Rats injected with the beta2-adrenoceptor agonist clenbuterol (2 mg kg-1 per day) for 18 days gained significantly more weight than controls. Tissue blood flow assessed 24 h after the last injection from the distribution of radiolabelled microspheres was increased in white (5 fold) and brown (3 fold) adipose tissue of clenbuterol-treated rats but was unaffected in kidney, brain and diaphragm, and was reduced by about 80% in skeletal muscle. Acute injection of clenbuterol one hour before measuring blood flow, increased blood flow to brown fat (20 fold) in both treated and control groups. Blood flow to skeletal muscle increased more in the rats treated chronically with clenbuterol (6 fold increase) than in control rats (2 fold increase), but absolute flow rates were still significantly lower in the rats treated chronically with clenbuterol. Skeletal muscle beta-adrenoceptor density and subtype were assessed from ligand binding and displacement studies using [3H]-dihydroalprenolol. Rats treated with clenbuterol for 18 days showed a 50% reduction in beta-receptor density, but the ratio of beta 1/beta 2-receptors was unaffected (15% beta 1/85% beta 2). The results indicate that, although clenbuterol produces acute increases in muscle blood flow, chronic treatment results in lower flow rates immediately (1 h) and 24 h after the previous injection. The attenuated response following chronic treatment is associated with a marked reduction in skeletal muscle beta-adrenoceptor density. The data suggest that any anabolic effects of clenbuterol on muscle which may require, or may be mediated by increases in blood supply, cannot be sustained by chronic treatment. Conversely, blood flow to white and brown adipose tissue would appear to be potentiated by chronic treatment, possibly reflecting increases in lipolytic and/or thermogenic activity.


A balance of outward and linear inward ionic currents is required for generation of slow-wave oscillations.
Regenerative inward currents help produce slow oscillations through a negative-slope conductance region of their current-voltage relationship that is well approximated by a linear negative conductance. We used dynamic-clamp injections of a linear current with such conductance, INL, to explore why some neurons can generate intrinsic slow oscillations whereas others cannot. We addressed this question in synaptically isolated neurons of the crab Cancer borealis after blocking action potentials. The pyloric network consists of a distinct pacemaker and follower neurons, all of which express the same complement of ionic currents. When the pyloric dilator (PD) neuron, a member of the pacemaker group, was injected with INL with dynamic clamp, it consistently produced slow oscillations. In contrast, all follower neurons failed to oscillate with INL To understand these distinct behaviors, we compared outward current levels of PD with those of follower lateral pyloric (LP) and ventral pyloric (VD) neurons. We found that LP and VD neurons had significantly larger high-threshold potassium currents (IHTK) than PD and LP had lower-transient potassium current (IA). Reducing IHTK pharmacologically enabled both LP and VD neurons to produce INL-induced oscillations, whereas modifying IA levels did not affect INL-induced oscillations. Using phase-plane and bifurcation analysis of a simplified model cell, we demonstrate that large levels of IHTK can block INL-induced oscillatory activity whereas generation of oscillations is almost independent of IA levels. These results demonstrate the general importance of a balance between inward pacemaking currents and high-threshold K+ current levels in determining slow oscillatory activity.NEW & NOTEWORTHY Pacemaker neuron-generated rhythmic activity requires the activation of at least one inward and one outward current. We have previously shown that the inward current can be a linear current (with negative conductance). Using this simple mechanism, here we demonstrate that the inward current conductance must be in relative balance with the outward current conductances to generate oscillatory activity. Surprisingly, an excess of outward conductances completely precludes the possibility of achieving such a balance.