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1.  U.S. National Institutes of Health Core Consolidation–Investing in Greater Efficiency 
The U.S. National Institutes of Health (NIH) invests substantial resources in core research facilities (cores) that support research by providing advanced technologies and scientific and technical expertise as a shared resource. In 2010, the NIH issued an initiative to consolidate multiple core facilities into a single, more efficient core. Twenty-six institutions were awarded supplements to consolidate a number of similar core facilities. Although this approach may not work for all core settings, this effort resulted in consolidated cores that were more efficient and of greater benefit to investigators. The improvements in core operations resulted in both increased services and more core users through installation of advanced instrumentation, access to higher levels of management expertise; integration of information management and data systems; and consolidation of billing; purchasing, scheduling, and tracking services. Cost recovery to support core operations also benefitted from the consolidation effort, in some cases severalfold. In conclusion, this program of core consolidation resulted in improvements in the effective operation of core facilities, benefiting both investigators and their supporting institutions.
doi:10.7171/jbt.15-2601-003
PMCID: PMC4310223  PMID: 25649473
centralized resources; infrastructure; shared technologies
2.  Adsorbed Fibrinogen Enhances Production of Bone- and Angiogenic-Related Factors by Monocytes/Macrophages 
Tissue Engineering. Part A  2013;20(1-2):250-263.
Macrophages are phagocytic cells with great importance in guiding multiple stages of inflammation and tissue repair. By producing a large number of biologically active molecules, they can affect the behavior of other cells and events, such as the foreign body response and angiogenesis. Since protein adsorption to biomaterials is crucial for the inflammatory process, we addressed the ability of the pro-inflammatory molecule fibrinogen (Fg) to modulate macrophage behavior toward tissue repair/regeneration. For this purpose, we used chitosan (Ch) as a substrate for Fg adsorption. Freshly isolated human monocytes were seeded on Ch substrates alone or previously adsorbed with Fg, and allowed to differentiate into macrophages for 10 days. Cell adhesion and morphology, formation of foreign body giant cells (FBGC), and secretion of a total of 80 cytokines and growth factors were evaluated. Both substrates showed similar numbers of adherent macrophages along differentiation as compared with RGD-coated surfaces, which were used as positive controls. Fg did not potentiate FBGC formation. In addition, actin cytoskeleton staining revealed the presence of punctuate F-actin with more elongated and interconnecting cells on Ch substrates. Antibody array screening and quantification of inflammation- and wound-healing-related factors indicated an overall reduction in Ch-based substrates versus RGD-coated surfaces. At late times, most inflammatory agents were down-regulated in the presence of Fg, in contrast to growth factor production, which was stimulated by Fg. Importantly, on Ch+Fg substrates, fully differentiated macrophages produced significant amounts of macrophage inflammatory protein-1delta (MIP-1δ), platelet-derived growth factor-BB, bone morphogenetic protein (BMP)-5, and BMP-7 compared with Ch alone. In addition, other important factors involved in bone homeostasis and wound healing, such as growth hormone, transforming growth factor-β3, and insulin-like growth factor-binding proteins, as well as several angiogenic mediators, including endocrine gland-derived vascular endothelial factor, fibroblast growth factor-7, and placental growth factor, were significantly promoted by Fg. This work provides a new perspective on the inflammatory response in the context of bone repair/regeneration mediated by a pro-inflammatory protein (Fg) adsorbed onto a biomaterial (Ch) that does not otherwise exhibit osteogenic properties.
doi:10.1089/ten.tea.2012.0439
PMCID: PMC3875152  PMID: 23937279
3.  Stratospheric controlled perturbation experiment: a small-scale experiment to improve understanding of the risks of solar geoengineering 
Although solar radiation management (SRM) through stratospheric aerosol methods has the potential to mitigate impacts of climate change, our current knowledge of stratospheric processes suggests that these methods may entail significant risks. In addition to the risks associated with current knowledge, the possibility of ‘unknown unknowns’ exists that could significantly alter the risk assessment relative to our current understanding. While laboratory experimentation can improve the current state of knowledge and atmospheric models can assess large-scale climate response, they cannot capture possible unknown chemistry or represent the full range of interactive atmospheric chemical physics. Small-scale, in situ experimentation under well-regulated circumstances can begin to remove some of these uncertainties. This experiment—provisionally titled the stratospheric controlled perturbation experiment—is under development and will only proceed with transparent and predominantly governmental funding and independent risk assessment. We describe the scientific and technical foundation for performing, under external oversight, small-scale experiments to quantify the risks posed by SRM to activation of halogen species and subsequent erosion of stratospheric ozone. The paper's scope includes selection of the measurement platform, relevant aspects of stratospheric meteorology, operational considerations and instrument design and engineering.
doi:10.1098/rsta.2014.0059
PMCID: PMC4240955  PMID: 25404681
geoengineering; solar radiation management; stratosphere; balloon; ozone depletion
4.  A novel algorithm for simplification of complex gene classifiers in cancer 
Cancer research  2013;73(18):5625-5632.
The clinical application of complex molecular classifiers as diagnostic or prognostic tools has been limited by the time and cost needed to apply them to patients. Using an existing fifty-gene expression signature known to separate two molecular subtypes of the pediatric cancer rhabdomyosarcoma, we show that an exhaustive iterative search algorithm can distill this complex classifier down to two or three features with equal discrimination. We validated the two-gene signatures using three separate and distinct data sets, including one that uses degraded RNA extracted from formalin-fixed, paraffin-embedded material. Finally, to demonstrate the generalizability of our algorithm, we applied it to a lung cancer data set to find minimal gene signatures that can distinguish survival. Our approach can easily be generalized and coupled to existing technical platforms to facilitate the discovery of simplified signatures that are ready for routine clinical use.
doi:10.1158/0008-5472.CAN-13-0324
PMCID: PMC4233649  PMID: 23913937
5.  Vitronectin is A Critical Protein Adhesion Substrate for IL-4-INDUCED Foreign Body Giant Cell Formation 
An in vitro system of interleukin (IL)-4-induced foreign body giant cell (FBGC) formation was utilized to define the adhesion protein substrate(s) that promotes this aspect of the foreign body reaction on biomedical polymers. Human monocytes were cultured on cell culture polystyrene surfaces that had been pre-adsorbed with a synthetic arginine-glycine-aspartate (RGD) peptide previously found to support optimal FBGC formation, or with various concentrations of potential physiological protein substrates, i.e. complement C3bi, collagen types I or IV, fibrinogen, plasma fibronectin, fibroblast fibronectin, laminin, thrombospondin, vitronectin, or von Willebrand factor. Cultures were evaluated on days 0 (1.5 hr), 3, and 7 by May-Grünwald/Giemsa staining. Initial monocyte adhesion occurred on all adsorbed proteins. However, by day 7 of culture, only vitronectin was striking in its ability to support significant macrophage adhesion, development, and fusion leading to FBGC formation. Vitronectin supported high degrees of FBGC formation at an absorption concentration between 5 and 25 μg per ml. These findings suggest that adsorbed vitronectin is critical in the collective events that support and promote FBGC formation on biomedical polymers, and that the propensity for vitronectin adsorption may underlie the material surface chemistry dependency of FBGC formation.
doi:10.1002/jbm.a.31658
PMCID: PMC4227597  PMID: 17994558
adhesion; integrins; monocyte; macrophage fusion; biomedical polymers
6.  Retinal Connectomics: Towards Complete, Accurate Networks 
Connectomics is a strategy for mapping complex neural networks based on high-speed automated electron optical imaging, computational assembly of neural data volumes, web-based navigational tools to explore 1012–1015 byte (terabyte to petabyte) image volumes, and annotation and markup tools to convert images into rich networks with cellular metadata. These collections of network data and associated metadata, analyzed using tools from graph theory and classification theory, can be merged with classical systems theory, giving a more completely parameterized view of how biologic information processing systems are implemented in retina and brain. Networks have two separable features: topology and connection attributes. The first findings from connectomics strongly validate the idea that the topologies complete retinal networks are far more complex than the simple schematics that emerged from classical anatomy. In particular, connectomics has permitted an aggressive refactoring of the retinal inner plexiform layer, demonstrating that network function cannot be simply inferred from stratification; exposing the complex geometric rules for inserting different cells into a shared network; revealing unexpected bidirectional signaling pathways between mammalian rod and cone systems; documenting selective feedforward systems, novel candidate signaling architectures, new coupling motifs, and the highly complex architecture of the mammalian AII amacrine cell. This is but the beginning, as the underlying principles of connectomics are readily transferrable to non-neural cell complexes and provide new contexts for assessing intercellular communication.
doi:10.1016/j.preteyeres.2013.08.002
PMCID: PMC4045117  PMID: 24016532
retina; neurons; connectome; networks; synapses; gap junctions
7.  TP53 Mutational Status Is a Potential Marker for Risk Stratification in Wilms Tumour with Diffuse Anaplasia 
PLoS ONE  2014;9(10):e109924.
Purpose
The presence of diffuse anaplasia in Wilms tumours (DAWT) is associated with TP53 mutations and poor outcome. As patients receive intensified treatment, we sought to identify whether TP53 mutational status confers additional prognostic information.
Patients and Methods
We studied 40 patients with DAWT with anaplasia in the tissue from which DNA was extracted and analysed for TP53 mutations and 17p loss. The majority of cases were profiled by copy number (n = 32) and gene expression (n = 36) arrays. TP53 mutational status was correlated with patient event-free and overall survival, genomic copy number instability and gene expression profiling.
Results
From the 40 cases, 22 (55%) had TP53 mutations (2 detected only after deep-sequencing), 20 of which also had 17p loss (91%); 18 (45%) cases had no detectable mutation but three had 17p loss. Tumours with TP53 mutations and/or 17p loss (n = 25) had an increased risk of recurrence as a first event (p = 0.03, hazard ratio (HR), 3.89; 95% confidence interval (CI), 1.26–16.0) and death (p = 0.04, HR, 4.95; 95% CI, 1.36–31.7) compared to tumours lacking TP53 abnormalities. DAWT carrying TP53 mutations showed increased copy number alterations compared to those with wild-type, suggesting a more unstable genome (p = 0.03). These tumours showed deregulation of genes associated with cell cycle and DNA repair biological processes.
Conclusion
This study provides evidence that TP53 mutational analysis improves risk stratification in DAWT. This requires validation in an independent cohort before clinical use as a biomarker.
doi:10.1371/journal.pone.0109924
PMCID: PMC4196953  PMID: 25313908
8.  Histologic and Clinical Characteristics Can Guide Staging Evaluations for Children and Adolescents With Rhabdomyosarcoma: A Report From the Children's Oncology Group Soft Tissue Sarcoma Committee 
Journal of Clinical Oncology  2013;31(26):3226-3232.
Purpose
To simplify the recommended staging evaluation by correlating tumor and clinical features with patterns of distant metastasis in newly diagnosed patients with embryonal rhabdomyosarcoma (ERMS) or alveolar rhabdomyosarcoma (ARMS).
Patients and Methods
Patient data from the Intergroup Rhabdomyosarcoma Study Group and the Children's Oncology Group over two periods were analyzed: 1991 to 1997 and 1999 to 2004. We used recursive partitioning analyses to identify factors (including histology, age, regional nodal and distant metastatic status, tumor size, local invasiveness, and primary site) that divided patients into subsets with the most different rates of metastatic disease.
Results
Of the 1,687 patients analyzed, 5.7% had lung metastases, 4.8% had bone involvement, and 6% had bone marrow (BM) involvement. Rhabdomyosarcoma (RMS) without local invasion (T1) had a low rate of metastasis for all distant sites, especially ERMS (0% bone, 0% BM). ARMS with local invasion (T2) had a higher rate of metastasis for all distant sites (13% lung, 18% bone, 23% BM). ERMS, T2 also had a higher rate of metastatic lung involvement (9%). The likelihood of bone or BM involvement increased in the presence of lung metastases (41% with, 6% without). Regional nodal metastases (N1) predicted a high rate of metastasis in all distant sites (14% lung, 14% bone, 18% BM). A staging algorithm was developed.
Conclusion
Staging studies in childhood RMS can be tailored to patients' presenting characteristics. Bone marrow aspirate and biopsy and bone scan are unnecessary in at least one third of patients with RMS.
doi:10.1200/JCO.2012.44.6476
PMCID: PMC3757291  PMID: 23940218
9.  The resemblance and disparity of gene expression in dormant and non-dormant seeds and crown buds of leafy spurge (Euphorbia esula) 
BMC Plant Biology  2014;14(1):216.
Background
Leafy spurge (Euphorbia esula L.) is a herbaceous perennial weed and dormancy in both buds and seeds is an important survival mechanism. Bud dormancy in leafy spurge exhibits three well-defined phases of para-, endo- and ecodormancy; however, seed dormancy for leafy spurge is classified as physiological dormancy that requires after-ripening and alternating temperature for maximal germination. Overlaps in transcriptome profiles between different phases of bud and seed dormancy have not been determined. Thus, we compared various phases of dormancy between seeds and buds to identify common genes and molecular processes, which should provide new insights about common regulators of dormancy.
Results
Cluster analysis of expression profiles for 201 selected genes indicated bud and seed samples clustered separately. Direct comparisons between buds and seeds are additionally complicated since seeds incubated at a constant temperature of 20°C for 21 days (21d C) could be considered paradormant (Para) because seeds may be inhibited by endosperm-generated signals, or ecodormant (Eco) because seeds germinate after being subjected to alternating temperature of 20:30°C. Since direct comparisons in gene expression between buds and seeds were problematic, we instead examined commonalities in differentially-expressed genes associated with different phases of dormancy. Comparison between buds and seeds (‘Para to Endo buds’ and ‘21d C to 1d C seeds’), using endodormant buds (Endo) and dormant seeds (1d C) as common baselines, identified transcripts associated with cell cycle (HisH4), stress response/transcription factors (ICE2, ERFB4/ABR1), ABA and auxin response (ABA1, ARF1, IAA7, TFL1), carbohydrate/protein degradation (GAPDH_1), and transport (ABCB2). Comparison of transcript abundance for the ‘Eco to Endo buds’ and ‘21d C to 1d C seeds’ identified transcripts associated with ABA response (ATEM6), auxin response (ARF1), and cell cycle (HisH4). These results indicate that the physiological state of 21d C seeds is more analogous to paradormant buds than that of ecodormant buds.
Conclusion
Combined results indicate that common molecular mechanisms associated with dormancy transitions of buds and seeds involve processes associated with ABA and auxin signaling and transport, cell cycle, and AP2/ERF transcription factors or their up-stream regulators.
Electronic supplementary material
The online version of this article (doi:10.1186/s12870-014-0216-4) contains supplementary material, which is available to authorized users.
doi:10.1186/s12870-014-0216-4
PMCID: PMC4256794  PMID: 25112962
Leafy spurge; Bud dormancy; Seed dormancy; Gene expression; Hormones; Transcription factors
10.  Children’s Oncology Group’s 2013 Blueprint for Research: Renal Tumors 
Pediatric blood & cancer  2012;60(6):994-1000.
Renal malignancies are among the most prevalent pediatric cancers. The most common is favorable histology Wilms tumor (FHWT), which has 5-year overall survival exceeding 90%. Other pediatric renal malignancies, including anaplastic Wilms tumor, clear cell sarcoma, malignant rhabdoid tumor, and renal cell carcinoma, have less favorable outcomes. Recent clinical trials have identified gain of chromosome 1q as a prognostic marker for FHWT. Upcoming studies will evaluate therapy adjustments based on this and other novel biomarkers. For high-risk renal tumors, new treatment regimens will incorporate biological therapies. A research blueprint, viewed from the perspective of the Children’s Oncology Group, is presented.
doi:10.1002/pbc.24419
PMCID: PMC4127041  PMID: 23255438
Wilms tumor; renal cell carcinoma; malignant rhabdoid tumor; clear cell sarcoma
11.  Longitudinal Performance of a Surgically Implanted Neuroprosthesis for Lower Extremity Exercise, Standing, and Transfers after Spinal Cord Injury 
Objective
To investigate the longitudinal performance of a surgically implanted neuroprosthesis for lower extremity exercise, standing, and transfers after spinal cord injury.
Design
Case series.
Setting
Research or outpatient physical therapy departments of four academic hospitals.
Participants
15 subjects with thoracic or low-cervical level spinal cord injuries who had received the 8-channel neuroprosthesis for exercise and standing.
Interventions
After completing rehabilitation with the device, the subjects were discharged to unrestricted home use of the system. A series of assessments were performed before discharge and at a follow-up appointment approximately one year later.
Main Outcome Measure(s)
Neuroprosthesis usage, maximum standing time, body weight support, knee strength, knee fatigue index, electrode stability, and component survivability.
Results
Levels of maximum standing time, body weight support, knee strength, and knee fatigue index were not statistically different from discharge to follow-up (p > 0.05). Additionally, neuroprosthesis usage was consistent with subjects choosing to use the system on approximately half of the days during each monitoring period. Although the number of hours using the neuroprosthesis remained constant, subjects shifted their usage to more functional standing versus more maintenance exercise, suggesting that the subjects incorporated the neuroprosthesis into their lives. Safety and reliability of the system were demonstrated by electrode stability and a high component survivability rate (>90%).
Conclusions
This group of 15 subjects is the largest cohort of implanted lower extremity neurorprosthetic exercise and standing system users. The safety and efficiency data from this group, and acceptance of the neuroprosthesis as demonstrated by continued usage, indicate that future efforts towards commercialization of a similar device may be warranted.
doi:10.1016/j.apmr.2012.01.001
PMCID: PMC4111081  PMID: 22541312
Electrical Stimulation; Exercise; Neural Prostheses; Spinal Cord Injuries; Weight-Bearing
12.  Considerations in the design of clinical trials for pediatric acute lymphoblastic leukemia 
Clinical investigation  2013;3(9):10.4155/cli.13.71.
Acute lymphoblastic leukemia (ALL) is the most common pediatric malignancy. Although outcomes for children with ALL have improved dramatically over the last 50 years, ALL remains the leading cause of childhood cancer death. In addition, high-risk patient subsets can be identified with significantly inferior survival. In the current era of therapies directed at specific molecular targets, the use of conventional randomized Phase III trials to show benefit from a new treatment regimen may not be feasible when these biologically defined subsets are small. This review presents the traditional approaches to designing trials for children with ALL, as well as innovative approaches attempting to study the benefit of new treatments as reliably as possible for patient subsets with distinctive biological characteristics.
doi:10.4155/cli.13.71
PMCID: PMC3834963  PMID: 24273641
acute lymphoblastic leukemia; clinical trials; historical controls
13.  Hexa-Acylated Lipid A Is Required for Host Inflammatory Response to Neisseria gonorrhoeae in Experimental Gonorrhea 
Infection and Immunity  2014;82(1):184-192.
Neisseria gonorrhoeae causes gonorrhea, a sexually transmitted infection characterized by inflammation of the cervix or urethra. However, a significant subset of patients with N. gonorrhoeae remain asymptomatic, without evidence of localized inflammation. Inflammatory responses to N. gonorrhoeae are generated by host innate immune recognition of N. gonorrhoeae by several innate immune signaling pathways, including lipooligosaccharide (LOS) and other pathogen-derived molecules through activation of innate immune signaling systems, including toll-like receptor 4 (TLR4) and the interleukin-1β (IL-1β) processing complex known as the inflammasome. The lipooligosaccharide of N. gonorrhoeae has a hexa-acylated lipid A. N. gonorrhoeae strains that carry an inactivated msbB (also known as lpxL1) gene produce a penta-acylated lipid A and exhibit reduced biofilm formation, survival in epithelial cells, and induction of epithelial cell inflammatory signaling. We now show that msbB-deficient N. gonorrhoeae induces less inflammatory signaling in human monocytic cell lines and murine macrophages than the parent organism. The penta-acylated LOS exhibits reduced toll-like receptor 4 signaling but does not affect N. gonorrhoeae-mediated activation of the inflammasome. We demonstrate that N. gonorrhoeae msbB is dispensable for initiating and maintaining infection in a murine model of gonorrhea. Interestingly, infection with msbB-deficient N. gonorrhoeae is associated with less localized inflammation. Combined, these data suggest that TLR4-mediated recognition of N. gonorrhoeae LOS plays an important role in the pathogenesis of symptomatic gonorrhea infection and that alterations in lipid A biosynthesis may play a role in determining symptomatic and asymptomatic infections.
doi:10.1128/IAI.00890-13
PMCID: PMC3911856  PMID: 24126526
14.  2010 Panel on the Biomaterials Grand Challenges 
In 2009, the National Academy for Engineering issued the Grand Challenges for Engineering in the 21st Century comprised of 14 technical challenges that must be addressed to build a healthy, profitable, sustainable, and secure global community (http://www.engineeringchallenges.org). Although crucial, none of the NEA Grand Challenges adequately addressed the challenges that face the biomaterials community. In response to the NAE Grand Challenges, Monty Reichert of Duke University organized a panel entitled Grand Challenges in Biomaterials at the at the 2010 Society for Biomaterials Annual Meeting in Seattle. Six members of the National Academies—Buddy Ratner, James Anderson, Allan Hoffman, Art Coury, Cato Laurencin, and David Tirrell—were asked to propose a grand challenge to the audience that, if met, would significantly impact the future of biomaterials and medical devices. Successfully meeting these challenges will speed the 60-plus year transition from commodity, off-the-shelf biomaterials to bioengineered chemistries, and biomaterial devices that will significantly advance our ability to address patient needs and also to create new market opportunities.
doi:10.1002/jbm.a.32969
PMCID: PMC4076103  PMID: 21171147
biomaterial; grand challenges; biocompatibility testing; smart polymers; regenerative medicine
15.  Certain Adenylated Non-Coding RNAs, Including 5′ Leader Sequences of Primary MicroRNA Transcripts, Accumulate in Mouse Cells following Depletion of the RNA Helicase MTR4 
PLoS ONE  2014;9(6):e99430.
RNA surveillance plays an important role in posttranscriptional regulation. Seminal work in this field has largely focused on yeast as a model system, whereas exploration of RNA surveillance in mammals is only recently begun. The increased transcriptional complexity of mammalian systems provides a wider array of targets for RNA surveillance, and, while many questions remain unanswered, emerging data suggest the nuclear RNA surveillance machinery exhibits increased complexity as well. We have used a small interfering RNA in mouse N2A cells to target the homolog of a yeast protein that functions in RNA surveillance (Mtr4p). We used high-throughput sequencing of polyadenylated RNAs (PA-seq) to quantify the effects of the mMtr4 knockdown (KD) on RNA surveillance. We demonstrate that overall abundance of polyadenylated protein coding mRNAs is not affected, but several targets of RNA surveillance predicted from work in yeast accumulate as adenylated RNAs in the mMtr4KD. microRNAs are an added layer of transcriptional complexity not found in yeast. After Drosha cleavage separates the pre-miRNA from the microRNA's primary transcript, the byproducts of that transcript are generally thought to be degraded. We have identified the 5′ leading segments of pri-miRNAs as novel targets of mMtr4 dependent RNA surveillance.
doi:10.1371/journal.pone.0099430
PMCID: PMC4057207  PMID: 24926684
16.  Beta-propeller protein-associated neurodegeneration: a new X-linked dominant disorder with brain iron accumulation 
Brain  2013;136(6):1708-1717.
Neurodegenerative disorders with high iron in the basal ganglia encompass an expanding collection of single gene disorders collectively known as neurodegeneration with brain iron accumulation. These disorders can largely be distinguished from one another by their associated clinical and neuroimaging features. The aim of this study was to define the phenotype that is associated with mutations in WDR45, a new causative gene for neurodegeneration with brain iron accumulation located on the X chromosome. The study subjects consisted of WDR45 mutation-positive individuals identified after screening a large international cohort of patients with idiopathic neurodegeneration with brain iron accumulation. Their records were reviewed, including longitudinal clinical, laboratory and imaging data. Twenty-three mutation-positive subjects were identified (20 females). The natural history of their disease was remarkably uniform: global developmental delay in childhood and further regression in early adulthood with progressive dystonia, parkinsonism and dementia. Common early comorbidities included seizures, spasticity and disordered sleep. The symptoms of parkinsonism improved with l-DOPA; however, nearly all patients experienced early motor fluctuations that quickly progressed to disabling dyskinesias, warranting discontinuation of l-DOPA. Brain magnetic resonance imaging showed iron in the substantia nigra and globus pallidus, with a ‘halo’ of T1 hyperintense signal in the substantia nigra. All patients harboured de novo mutations in WDR45, encoding a beta-propeller protein postulated to play a role in autophagy. Beta-propeller protein-associated neurodegeneration, the only X-linked disorder of neurodegeneration with brain iron accumulation, is associated with de novo mutations in WDR45 and is recognizable by a unique combination of clinical, natural history and neuroimaging features.
doi:10.1093/brain/awt095
PMCID: PMC3673459  PMID: 23687123
iron; NBIA; autophagy; basal ganglia; Rett syndrome
17.  Improved synthesis of structural analogues of (−)-epicatechin gallate for modulation of staphylococcal β-lactam resistance☆ 
Tetrahedron  2014;70(21):3485-3490.
The high-yielding synthesis of enantiomerically pure epicatechin gallate analogues where the A and/or B-ring hydroxylation is reduced or altered has been achieved by optimising routes to the catechin stereochemistry. The B-ring analogues were synthesised by using an electrophilic ring closure onto an enantiomerically enriched epoxide as a key step. The A and B-ring hydroxyl-deleted analogues were synthesised through a Mitsunobu cyclisation. For the B-ring analogues, the anti- (catechin) stereochemistry was converted to the syn- (epicatechin) stereochemistry by a known oxidation/reduction protocol. Absolute stereochemistry was derived from either a Sharpless epoxidation or asymmetric dihydroxylation.
Graphical abstract
doi:10.1016/j.tet.2014.03.052
PMCID: PMC4006114  PMID: 24876661
(−)-Epicatechin gallate analogues; Asymmetric synthesis; Cyclisation; Mitsunobu; MRSA
18.  Mortality Increase in Late-Middle and Early-Old Age: Heterogeneity in Death Processes as a New Explanation 
Demography  2013;50(5):1563-1591.
Deviations from the Gompertz law of exponential mortality increases in late-middle and early-old age are commonly neglected in overall mortality analyses. In this study, we examined mortality increase patterns between ages 40 and 85 in 16 low-mortality countries and demonstrated sex differences in these patterns, which also changed across period and cohort. These results suggest that the interaction between aging and death is more complicated than what is usually assumed from the Gompertz law and also challenge existing biodemographic hypotheses about the origin and mechanisms of sex differences in mortality. We propose a two-mortality model that explains these patterns as the change in the composition of intrinsic and extrinsic death rates with age. We show that the age pattern of overall mortality and the population heterogeneity therein are possibly generated by multiple dynamics specified by a two-mortality model instead of a uniform process throughout most adult ages.
doi:10.1007/s13524-013-0222-4
PMCID: PMC4028711  PMID: 23743628
Mortality acceleration; Sex differences; Two-mortality processes; Vitality
19.  Instability of Self-Assembled Monolayers (SAM) as a Model Material System for Macrophage/FBGC Cellular Behavior 
Novel self-assembled monolayers (SAMs) designed to present homogenous surface chemistries were utilized to further investigate the material surface chemistry dependent macrophage and foreign body giant cell (FBGC) behaviors including macrophage adhesion, fusion, and apoptosis. Contact angle analysis revealed instabilities in the –CH3 and –COOH terminate SAM surfaces upon incubation in serum-free media at 37oC or under dry, room temperatureconditions. Further analysis indicated that the –CH3 terminated SAM surface degraded rapidly within 2 hours and loss of sufficient SAM units to be comparable to the gold (Au) control surface within 24 hours of incubation in serum-free media (SFM) at 37oC. After 5days of incubation in SFM at 37oC, the contact angles for the –COOH terminated SAMsurfaces increased markedly. AFM analysis confirmed the desorption of –CH3 terminated SAM molecules from the surface with increased roughness and marked appearance of peaks andvalleys within 2 hours. A decrease in the thickness of the –COOH terminated SAM surface also suggests molecular desorption over time. No significant changes in contact angle or AFM analyses were observed on the –OH terminated SAM surfaces. Cellular adhesion decreased morerapidly on the Au control and –CH3 terminated SAM surfaces in comparison to the other surfaces. However by day 10, cellular adhesion, fusion, and apoptosis were comparable on all SAM surfaces and the Au control . These studies suggest that SAM surfaces may not be suitable for long-term studies where material dependent properties are investigated.
doi:10.1002/jbm.a.31660
PMCID: PMC4017672  PMID: 18412136
self-assembled monolayers; instability; macrophage; foreign body giant cell; contact angles
20.  Patterns of Chemotherapy-Induced Toxicities in Younger Children and Adolescents with Rhabdomyosarcoma: A Report from the Children’s Oncology Group Soft Tissue Sarcoma Committee 
Cancer  2011;118(4):1130-1137.
Purpose
Patients aged older than 10 years with rhabdomyosarcoma (RMS) have an inferior outcome compared to patients aged 1–9 years which might be explained by toxicities (adverse events, AE) resulting in chemotherapy dose reductions.
Methods
AEs observed during one of 3 randomized chemotherapy regimens (VAC, VAI or VIE) in Intergroup Rhabdomyosarcoma Study (IRS)-IV were recorded. The incidence of toxicities by age and treatment regimen was determined. The odds of developing an AE in a particular age group (5–9, 10–14, and 15–20 years) were compared to the control age group, 1–4 years.
Results
657 patients were eligible for analysis. Estimated 5-year EFS were 78%, 83%, 67%, and 58% for age groups 1–4, 5–9, 10–14, and 15–20, respectively. Patients 15–20 years experienced less neutropenia (OR 0.43, p<0.0001), thrombocytopenia (OR 0.41, p<0.0001), anemia (OR 0.34, p<0.0001) and infection (OR 0.41, p<0.0001) compared to younger patients, despite receiving similar amounts of chemotherapy. In contrast, peripheral nervous system (PNS) toxicity was higher in adolescents older than 10 years (OR 4.18, p<0.0001). Females experienced more neutropenia (OR 1.28, p=0.05) and thrombocytopenia (OR 1.26, p=0.06) compared to males.
Conclusions
Adolescents receiving treatment for RMS experience significantly less hematologic and more PNS toxicity compared to younger children despite receiving similar amounts of chemotherapy. Although outcome is inferior in adolescents, it is unclear whether the differences in toxicity observed in our study impact on outcome. Future studies examining the age and gender-related differences in pharmacokinetics of chemotherapy are necessary.
doi:10.1002/cncr.26358
PMCID: PMC4008942  PMID: 21761400
adolescent; rhabdomyosarcoma; toxicity; chemotherapy; age; gender
21.  The immunophenotype of T-lymphoblastic lymphoma in children and adolescents: a Children’s Oncology Group report 
British journal of haematology  2012;159(4):454-461.
Summary
T-lymphoblastic leukaemia (T-ALL) and T-lymphoblastic lymphoma (T-LBL) are neoplasms derived from immature lymphoid cells of T-cell lineage. These neoplasms are biologically similar, but significant differences may exist between the two given their clinical differences. Although ample data regarding the immunophenotypic characterization T-ALL are available, there is a paucity of such data in children and adolescents with T-LBL. We used flow cytometry and/or immunohistochemistry to characterize the immunophenotypic profile of 180 children and adolescents with newly diagnosed T-LBL enrolled in the Children’s Oncology Group 5971 study. Multiple T-cell, B-cell, myeloid, and other markers were evaluated. We identified diagnostically useful immunophenotypic features of T-LBL as well as distinct immunophenotypic subgroups, although none of these was statistically related to event-free or overall survival in this retrospective analysis. Further studies of biologically and immunophenotypically distinct subgroups of T-LBL, such as the early T-cell precursor phenotype, are warranted.
doi:10.1111/bjh.12042
PMCID: PMC4008319  PMID: 22994934
T-lymphoblastic lymphoma; early T-cell precursor; immunophenotypic analysis; paediatric lymphoma; T-cell antigens
22.  Lymph node involvement in Wilms tumor: results from National Wilms Tumor Studies 4 and 5 
Journal of pediatric surgery  2012;47(4):700-706.
Purpose
The aim of the study was to determine the prognostic impact of lymph node (LN) involvement and sampling in patients with Wilms tumor (WT) and the minimum number of LNs needed for accurate staging.
Methods
We reviewed all patients with unilateral, nonmetastatic WT enrolled in the National Wilms Tumor Study 4 or 5. Data were abstracted on patient demographics, tumor histology, staging, number of LNs sampled, and disease-specific and overall patient outcomes.
Results
A total of 3409 patients had complete information on LN sampling. Five-year event-free survival (EFS) was lower in patients with nodal disease (P = .001); the effect of LN positivity was greater for patients with anaplastic (P = .047) than with favorable histology (P = .02). The likelihood of obtaining a positive LN was higher when sampling at least 7 LNs. However, after controlling for tumor histology and stage, the number of LNs sampled did not predict EFS variations (P = .75). Among patients with stage II disease, patients with LN sampling (P = .055) had improved EFS, largely reflecting poorer EFS in patients with anaplastic tumors (P = .03).
Conclusions
Lymph node sampling is particularly important for patients with stage II anaplastic WT. Although the likelihood of finding a positive LN was greater when more than 7 LNs were sampled, EFS was not impacted by the number of LNs sampled.
doi:10.1016/j.jpedsurg.2011.08.017
PMCID: PMC3976547  PMID: 22498384
Wilms tumor; Lymph node; Treatment outcome; Child
23.  Staphylococcal Phenotypes Induced by Naturally Occurring and Synthetic Membrane-Interactive Polyphenolic β-Lactam Resistance Modifiers 
PLoS ONE  2014;9(4):e93830.
Galloyl catechins, in particular (-)-epicatechin gallate (ECg), have the capacity to abrogate β-lactam resistance in methicillin-resistant strains of Staphylococcus aureus (MRSA); they also prevent biofilm formation, reduce the secretion of a large proportion of the exoproteome and induce profound changes to cell morphology. Current evidence suggests that these reversible phenotypic traits result from their intercalation into the bacterial cytoplasmic membrane. We have endeavoured to potentiate the capacity of ECg to modify the MRSA phenotype by stepwise removal of hydroxyl groups from the B-ring pharmacophore and the A:C fused ring system of the naturally occurring molecule. ECg binds rapidly to the membrane, inducing up-regulation of genes responsible for protection against cell wall stress and maintenance of membrane integrity and function. Studies with artificial membranes modelled on the lipid composition of the staphylococcal bilayer indicated that ECg adopts a position deep within the lipid palisade, eliciting major alterations in the thermotropic behaviour of the bilayer. The non-galloylated homolog (-)-epicatechin enhanced ECg-mediated effects by facilitating entry of ECg molecules into the membrane. ECg analogs with unnatural B-ring hydroxylation patterns induced higher levels of gene expression and more profound changes to MRSA membrane fluidity than ECg but adopted a more superficial location within the bilayer. ECg possessed a high affinity for the positively charged staphylococcal membrane and induced changes to the biophysical properties of the bilayer that are likely to account for its capacity to disperse the cell wall biosynthetic machinery responsible for β-lactam resistance. The ability to enhance these properties by chemical modification of ECg raises the possibility that more potent analogs could be developed for clinical evaluation.
doi:10.1371/journal.pone.0093830
PMCID: PMC3974817  PMID: 24699700
24.  ON Cone Bipolar Cell Axonal Synapses in the OFF Inner Plexiform Layer of the Rabbit Retina 
The Journal of comparative neurology  2013;521(5):977-1000.
Analysis of the rabbit retinal connectome RC1 reveals that the division between the ON and OFF inner plexiform layer (IPL) is not structurally absolute. ON cone bipolar cells make non-canonical axonal synapses onto specific targets and receive amacrine cell synapses in the nominal OFF layer, creating novel motifs, including inhibitory crossover networks. Automated transmission electron microscope (ATEM) imaging, molecular tagging, tracing, and rendering of ≈ 400 bipolar cells reveals axonal ribbons in 36% of ON cone bipolar cells, throughout the OFF IPL. The targets include GABA-positive amacrine cells (γACs), glycine-positive amacrine cells (GACs) and ganglion cells. Most ON cone bipolar cell axonal contacts target GACs driven by OFF cone bipolar cells, forming new architectures for generating ON-OFF amacrine cells. Many of these ON-OFF GACs target ON cone bipolar cell axons, ON γACs and/or ON-OFF ganglion cells, representing widespread mechanisms for OFF to ON crossover inhibition. Other targets include OFF γACs presynaptic to OFF bipolar cells, forming γAC-mediated crossover motifs. ON cone bipolar cell axonal ribbons drive bistratified ON-OFF ganglion cells in the OFF layer and provide ON drive to polarity-appropriate targets such as bistratified diving ganglion cells (bsdGCs). The targeting precision of ON cone bipolar cell axonal synapses shows that this drive incidence is necessarily a joint distribution of cone bipolar cell axonal frequency and target cell trajectories through a given volume of the OFF layer. Such joint distribution sampling is likely common when targets are sparser than sources and when sources are coupled, as are ON cone bipolar cells.
doi:10.1002/cne.23244
PMCID: PMC3560306  PMID: 23042441
retina; inner plexiform layer; connectomics; circuitry; neural network; bipolar cell; axonal ribbon; axonal cistern; amacrine cell; bistratified diving ganglion cell; intrinsically photosensitive ganglion cell; crossover inhibition; within channel inhibition; ON-OFF crosstalk; functional network; structure-function; network topology
25.  A high-resolution morphological and ultrastructural map of anterior sensory cilia and glia in Caenorhabditis elegans 
eLife  2014;3:e01948.
Many primary sensory cilia exhibit unique architectures that are critical for transduction of specific sensory stimuli. Although basic ciliogenic mechanisms are well described, how complex ciliary structures are generated remains unclear. Seminal work performed several decades ago provided an initial but incomplete description of diverse sensory cilia morphologies in C. elegans. To begin to explore the mechanisms that generate these remarkably complex structures, we have taken advantage of advances in electron microscopy and tomography, and reconstructed three-dimensional structures of fifty of sixty sensory cilia in the C. elegans adult hermaphrodite at high resolution. We characterize novel axonemal microtubule organization patterns, clarify structural features at the ciliary base, describe new aspects of cilia–glia interactions, and identify structures suggesting novel mechanisms of ciliary protein trafficking. This complete ultrastructural description of diverse cilia in C. elegans provides the foundation for investigations into underlying ciliogenic pathways, as well as contributions of defined ciliary structures to specific neuronal functions.
DOI: http://dx.doi.org/10.7554/eLife.01948.001
eLife digest
To survive, animals must constantly gather information about their surroundings and then decide how to respond. Animals rely on cells called sensory neurons to help them perceive and process this information, and these neurons in most animals have smaller structures called cilia that help them to gather this information. The structures of these cilia can range from simple hair-like rods to complex branched arbors. Defective cilia can lead to cell degeneration and death.
Scientists have identified and determined the functions of many of the 60 sensory neurons with cilia in C. elegans, a tiny roundworm with a simple nervous system. These experiments have revealed that the shapes of these cilia are quite diverse, and that the shape determines the type of information the neurons process. Learning more about how cilia are shaped, and how these shapes allow them to perform specific sensory functions, would give scientists a better understanding of how the brain processes sensory information.
Doroquez et al. have now taken advantage of advances in imaging technology to generate highly detailed three-dimensional reconstructions of the cilia on 50 neurons in the nose of C. elegans. The experiments involved rapidly freezing the worms, slowly replacing the frozen water molecules with a preservative solution, and then embedding in resin. This allowed Doroquez et al. to slice the samples into very thin sections—some 1400 times thinner than a sheet of paper—and then image them with transmission electron microscopy and electron tomography. Finally, all these images were combined in a computer to produce 3D models of the cilia.
The models reveal a wide range of cilia structures, including some that have never been examined in detail before. Doroquez et al. were also able to see detailed structures within the cilia, including compartments that determine which proteins should enter into, or be excluded from, an individual cilium. The models, along with the results of previous studies, suggest that cilia are shaped by genetic factors and also by interactions with the environment. This detailed description of diverse cilia structures should now allow researchers to identify the genes that determine their unique shapes, and explore how specific shapes contribute to specific sensory functions.
DOI: http://dx.doi.org/10.7554/eLife.01948.002
doi:10.7554/eLife.01948
PMCID: PMC3965213  PMID: 24668170
cilia; electron microscopy; electron tomography; C. elegans

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