NIH Music-Based Intervention Toolkit: Music-Based Interventions for Brain Disorders of Aging.

Music-based interventions (MBIs) show promise for managing symptoms of various brain disorders. To fully realize the potential of MBIs and dispel the outdated misconception that MBIs are rooted in soft science, the NIH is promoting rigorously designed, well-powered MBI clinical trials. The pressing need of guidelines for scientifically rigorous studies with enhanced data collection brought together the Renée Fleming Foundation, the Foundation for the NIH, the Trans-NIH Music and Health Working Group, and an interdisciplinary scientific expert panel to create the NIH MBI Toolkit for research on music and health across the lifespan. The Toolkit defines the building blocks of MBIs, including a consolidated set of common data elements for MBI protocols, and core datasets of outcome measures and biomarkers for brain disorders of aging that researchers may select for their studies. Utilization of the guiding principles in this Toolkit will be strongly recommended for NIH-funded studies of MBIs.

Development of Common Data Elements for Use in Chiari Malformation Type I Clinical Research: An NIH/NINDS Project.

The management of Chiari I malformation (CMI) is controversial because treatment methods vary and treatment decisions rest on incomplete understanding of its complex symptom patterns, etiologies, and natural history. Validity of studies that attempt to compare treatment of CMI has been limited because of variable terminology and methods used to describe study subjects. The goal of this project was to standardize terminology and methods by developing a comprehensive set of Common Data Elements (CDEs), data definitions, case report forms (CRFs), and outcome measure recommendations for use in CMI clinical research, as part of the CDE project at the National Institute of Neurological Disorders and Stroke (NINDS) of the US National Institutes of Health. A working group, comprising over 30 experts, developed and identified CDEs, template CRFs, data dictionaries, and guidelines to aid investigators starting and conducting CMI clinical research studies. The recommendations were compiled, internally reviewed, and posted online for external public comment. In October 2016, version 1.0 of the CMI CDE recommendations became available on the NINDS CDE website. The recommendations span these domains: Core Demographics/Epidemiology; Presentation/Symptoms; Co-Morbidities/Genetics; Imaging; Treatment; and Outcome. Widespread use of CDEs could facilitate CMI clinical research trial design, data sharing, retrospective analyses, and consistent data sharing between CMI investigators around the world. Updating of CDEs will be necessary to keep them relevant and applicable to evolving research goals for understanding CMI and its treatment.

Performance of two respiratory protective devices used by home-attending health care workers (a pilot study).

OBJECTIVES: This pilot study aimed at determining the Workplace Protection Factor (WPF) for respiratory protective devices widely used by health care workers to reduce exposure to potentially hazardous aerosols when attending patients in their homes. Two devices were tested, an N95 filtering facepiece respirator (FFR) and a surgical mask (SM). METHODS: Three home-attending health care workers were recruited, medically cleared and fit tested. At the workplace, the aerosol concentrations outside (Cout) and inside (Cin) of the tested respiratory protective device worn by a subject were measured using two simultaneously operating P-Trak condensation particle counters within the particle size range of approximately 20-1,000 nm. Real-time and integrated (time-weighted average, TWA) values of WPF = Cout/Cin were determined. RESULTS: This pilot study demonstrated that the WPF of the tested N95 FFR consistently exceeded that of the SM. The WPFTWA(C) values calculated for the entire test time (based on the TWA aerosol concentration values) ranged from 29 to 40 and 2 to 9, respectively. In all cases, the N95 FFR provided protection above the Occupational Safety and Health Administration's (OSHA) assigned protection factor of 10, whereas the SM often offered little or essentially no protection against the measured sub-micrometer aerosol particles. For both devices, the protection level was found to depend on activity. For example, the WPFTWA(C) for one subject wearing the N95 FFR was 56 during normal activity but fell almost 70% during tracheal suctioning. It is explicable considering that different procedures implemented by health care workers in homes generate particles of different sizes and require different body movements; both factors are anticipated to affect the WPF. CONCLUSIONS: Wearing an N95-certified respirator helps significantly reduce the aerosol inhalation exposure of home-attending health care workers. An SM offers much lower protection. The WPF depends on several factors, including, but not limited to, the health care worker's activity and/or body movements; the WPF varies from one worker to another.

Zebrafish Lmx1b.1 and Lmx1b.2 are required for maintenance of the isthmic organizer.

The mesencephalic and metencephalic region (MMR) of the vertebrate central nervous system develops in response to signals produced by the isthmic organizer (IsO). We have previously reported that the LIM homeobox transcription factor Lmx1b is expressed within the chick IsO, where it is sufficient to maintain expression of the secreted factor wnt1. In this paper, we show that zebrafish express two Lmx1b orthologs, lmx1b.1 and lmx1b.2, in the rostral IsO, and demonstrate that these genes are necessary for key aspects of MMR development. Simultaneous knockdown of Lmx1b.1 and Lmx1b.2 using morpholino antisense oligos results in a loss of wnt1, wnt3a, wnt10b, pax8 and fgf8 expression at the IsO, leading ultimately to programmed cell death and the loss of the isthmic constriction and cerebellum. Single morpholino knockdown of either Lmx1b.1 or Lmx1b.2 has no discernible effect on MMR development. Maintenance of lmx1b.1 and lmx1b.2 expression at the isthmus requires the function of no isthmus/pax2.1, as well as Fgf signaling. Transient misexpression of Lmx1b.1 or Lmx1b.2 during early MMR development induces ectopic wnt1 and fgf8 expression in the MMR, as well as throughout much of the embryo. We propose that Lmx1b.1- and Lmx1b.2-mediated regulation of wnt1, wnt3a, wnt10b, pax8 and fgf8 maintains cell survival in the isthmocerebellar region.

Purification and characterization of 2'aminobiphenyl-2,3-diol 1,2-dioxygenase from Pseudomonas sp. LD2.

Carbazole is a nitrogen-containing heteroaromatic compound that occurs as a widespread and mutagenic environmental pollutant. The 2'aminobiphenyl-2,3-diol 1,2-dioxygenase involved in carbazole degradation was purified to near electrophoretic homogeneity from Pseudomonas sp. LD2 by a combination of ion-exchange chromatography, ammonium sulfate precipitation, and hydrophobic interaction chromatography. This purification was challenging due to the great instability of the enzyme under many standard conditions. The enzyme was also purified to electrophoretic homogeneity from recombinant Escherichia coli expressing the 2'aminobiphenyl-2,3-diol 1,2-dioxygenase-encoding gene cloned from Pseudomonas sp. LD2. The molecular mass of the native enzyme was determined by gel filtration to be 70 kDa. The subunit molecular masses were determined to be 25 and 8 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, indicating that the dioxygenase is an [alpha2beta2] heterotetramer. The optimal temperature and pH for the enzymatic production of 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid (HOPDA) from 2,3-dihydroxybiphenyl were determined to be 40 degrees C and 8.0, respectively. The maximum observed specific activity on 2,3-dihydroxybiphenyl was 48.1 mmol HOPDA min(-1) mg(-1). This indicated a maximum observed turnover rate of 360,000 molecules HOPDA enz(-1) s(-1). The K'm inhibition constant Ks and Vmax on 2,3 dihydroxybiphenyl were determined to be 5 microM, 37 microM, and 44 mmol min(-1) mg(-1), respectively. These results show that 2'aminobiphenyl-2,3-diol 1,2-dioxygenase is a meta-cleavage enzyme related to the 4,5-protocatechuate dioxygenase family, with comparable purification challenges posed by intrinsic enzyme instability.

cBlimp-1 expression in chick limb bud development.

Blimp is a transcriptional repressor that has been identified in humans, mice and frogs. Here, we describe the cloning of the gene that encodes for the chick homolog of Blimp, cBlimp-1, and the tempo-spatial expression pattern of cBlimp-1, during chick development. cBlimp-1 whose gene product shares a high level of identity with its homologs from other organisms, was found to be expressed in the apical ectodermal ridge and posterior dorsal ectoderm of developing limb buds. In addition, cBlimp-1 is expressed in the developing eyes, branchial arches and otic placodes.

Purification and properties of 2-hydroxy-6-oxo-6-(2'-aminophenyl)hexa-2,4-dienoic acid hydrolase involved in microbial degradation of carbazole.

Hydrolysis following meta-ring cleavage by a dioxygenase is a well-known step in aromatic compound metabolism. The 2-hydroxy-6-oxo-6-(2'-aminophenyl)hexa-2,4-dienoic acid hydrolase from Pseudomonas LD2 is a new member of the small group of characterized aromatic hydrolases that catalyze the cleavage of C-C bonds. In this study, the His(6)-tagged 2-hydroxy-6-oxo-6-(2'-aminophenyl)hexa-2,4-dienoic acid (HOPDA) hydrolase was purified from a recombinant Escherichia coli strain utilizing immobilized metal affinity chromatography. 2-Hydroxy-6-oxo-6-(2'-aminophenyl)hexa-2,4-dienoic acid hydrolase is a colorless homodimer with no cofactor requirement. The enzyme actively converted HOPDA into benzoic acid and 2-hydroxypenta-2,4-dienoic acid. The enzyme exhibited activity between pH 6.5 and 10.5 with a maximum activity at pH 7.0. The optimum temperature at pH 7.0 was 60 degrees C. The calculated K'(m) for HOPDA was 4.6 microM, the V(max) was 3.3 micromol min(-1), and the K(s) was 70.0 microM. This corresponds to a maximum specific turnover rate of 1300 HOPDAs(-1)dimer(-1). The deduced amino acid sequence of CarC showed 30.3, 31.3, and 31.8% identity with TodF (P. putida F1), XylF (P. putida), and DmpD (Pseudomonas sp. CF600), respectively, which are meta-cleavage compound hydrolases from other Pseudomonads. The amino acid sequence Gly-X-Ser-X-Gly, which is highly conserved in these hydrolases, is also found in CarC. Lysates from a strain expressing enzyme in which the putative active site serine is mutated to alanine showed a significant reduction in activity.

Recombinant carbazole-degrading strains for enhanced petroleum processing.

Biotechnological upgrading of fossil fuels is of increasing interest as remaining stocks of petroleum show increasing levels of contaminants such as heavy metals, sulfur and nitrogen-containing heteroaromatic compounds. Carbazole is of particular interest as a major petroleum component known to reduce refining yields through catalyst poisoning. In this study, the biotransformation of carbazole was successfully demonstrated in a liquid two-phase system, when solubilized in either 1-methylnaphthalene or in diesel fuel. The effects of solvent toxicity were investigated by expressing the carbazole-transformation genes from MB1332, a rifampicin-resistant derivative of Pseudomonas sp. LD2, in a solvent-resistant heterologous host, P. putida Idaho [1]. This solvent-resistant strain successfully degraded carbazole solubilized in 1-methylnaphthalene and in the presence of 10 vol% xylenes similar to the non-recombinant strain Pseudomonas sp. LD2. Identification of a suitable recombinant host, however, was essential for further investigations of partial pathway transformations. Recombinant P. putida Idaho expressing only the initial dioxygenase enzymes transformed carbazole to an intermediate well retained in the oil phase. Partial carbazole transformation converts carbazole to non-aromatic species; their effect is unknown on refinery catalyst poisoning, but would allow almost complete retention of carbon content and fuel value.

Making connections: the development of mesencephalic dopaminergic neurons.

The disorders of two adjacent sets of mesencephalic dopaminergic (MDNs) are associated with two significant health problems: Parkinson's disease and drug addiction. Because of this, a great deal of research has focused on understanding the growth, development and maintenance of MDNs. Many transcription factors and signaling pathways are known to be required for normal MDNs formation, but a unified model of MDN development is still unclear. The long-term goal is to design therapeutic strategies to: (i) nurture and/or heal endogenous MDNs, (ii) replace the affected tissue with exogenous MDNs from in vitro cultivated stem cells and (iii) restore normal connectivity. Recent developmental biology studies show great promise in understanding how MDNs develop both in vivo and in vitro. This information has great therapeutic value and may provide insight into how environmental and genetic factors increase vulnerability to addiction.