Land use mix and physical activity in middle-aged and older adults: a longitudinal study examining changes in land use mix in two Dutch cohorts.

BACKGROUND: With urbanization and aging increasing in coming decades, societies face the challenge of keeping aging populations active. Land use mix (LUM) has been associated with cycling and walking, but whether changes in LUM relate to changes in cycling/walking is less known. OBJECTIVES: Our objective was to study the effect of LUM on cycling/walking in two Dutch aging cohorts using data with 10 years of follow-up. METHODS: Data from 1183 respondents from the Health and Living Conditions of the Population of Eindhoven and Surroundings (GLOBE) study and 918 respondents from the Longitudinal Aging Study Amsterdam (LASA) were linked to LUM in 1000-m sausage network buffers at three time-points. Cycling/walking outcomes were harmonized to include average minutes spent cycling/walking per week. Data was pooled and limited to respondents that did not relocate between follow-up waves. Associations between LUM and cycling/walking were estimated using a Random Effects Within-Between (REWB) model that allows for the estimation of both within and between effects. Sensitivity analyses were performed on smaller (500-m) and larger (1600-m) buffers. RESULTS: We found evidence of between-individual associations of LUM in 1000-m buffers and walking (ß: 11.10, 95% CI: 0.08; 21.12), but no evidence of within-associations in 1000-m buffers. Sensitivity analyses using 500-m buffers showed similar between-associations, but negative within-associations (ß: -35.67, 95% CI: - 68.85; - 2.49). We did not find evidence of between-individual associations of LUM in any buffer size and cycling, but did find evidence of negative within-associations between LUM in 1600-m buffers and cycling (ß: -7.49, 95% CI: - 14.31; - 0.66). DISCUSSION: Our study found evidence of positive associations between LUM and average walking time, but also some evidence of negative associations between a change in LUM and cycling/walking. LUM appears to be related to cycling/walking, but the effect of changes in LUM on cycling/walking is unclear.

Associations of neighbourhood sociodemographic characteristics with depressive and anxiety symptoms in older age: Results from a 5-wave study over 15 years.

We examined the long-term association between objective neighbourhood sociodemographic characteristics (index of socioeconomic position (SEP), average income, percent low-income earners, average house price, percent immigrants and urban density) with depressive and anxiety symptoms, covering five 3-year waves of the Longitudinal Aging Study Amsterdam (n = 3,772). Multi-level regression models assessed each neighbourhood-level characteristic separately, adjusting for individual-level covariates. A higher percentage of immigrants and higher urban density, but not other neighbourhood characteristics, were significantly associated with depressive and anxiety symptoms over time in models adjusted for individual SEP. Results of time interaction models indicated that the associations were stable over the 15-year period.

Tumor-Specific Mitochondrial DNA Variants Are Rarely Detected in Cell-Free DNA.

The use of blood-circulating cell-free DNA (cfDNA) as a "liquid biopsy" in oncology is being explored for its potential as a cancer biomarker. Mitochondria contain their own circular genomic entity (mitochondrial DNA, mtDNA), up to even thousands of copies per cell. The mutation rate of mtDNA is several orders of magnitude higher than that of the nuclear DNA. Tumor-specific variants have been identified in tumors along the entire mtDNA, and their number varies among and within tumors. The high mtDNA copy number per cell and the high mtDNA mutation rate make it worthwhile to explore the potential of tumor-specific cf-mtDNA variants as cancer marker in the blood of cancer patients. We used single-molecule real-time (SMRT) sequencing to profile the entire mtDNA of 19 tissue specimens (primary tumor and/or metastatic sites, and tumor-adjacent normal tissue) and 9 cfDNA samples, originating from 8 cancer patients (5 breast, 3 colon). For each patient, tumor-specific mtDNA variants were detected and traced in cfDNA by SMRT sequencing and/or digital PCR to explore their feasibility as cancer biomarker. As a reference, we measured other blood-circulating biomarkers for these patients, including driver mutations in nuclear-encoded cfDNA and cancer-antigen levels or circulating tumor cells. Four of the 24 (17%) tumor-specific mtDNA variants were detected in cfDNA, however at much lower allele frequencies compared to mutations in nuclear-encoded driver genes in the same samples. Also, extensive heterogeneity was observed among the heteroplasmic mtDNA variants present in an individual. We conclude that there is limited value in tracing tumor-specific mtDNA variants in blood-circulating cfDNA with the current methods available.

Sensitive detection of mitochondrial DNA variants for analysis of mitochondrial DNA-enriched extracts from frozen tumor tissue.

Large variation exists in mitochondrial DNA (mtDNA) not only between but also within individuals. Also in human cancer, tumor-specific mtDNA variation exists. In this work, we describe the comparison of four methods to extract mtDNA as pure as possible from frozen tumor tissue. Also, three state-of-the-art methods for sensitive detection of mtDNA variants were evaluated. The main aim was to develop a procedure to detect low-frequent single-nucleotide mtDNA-specific variants in frozen tumor tissue. We show that of the methods evaluated, DNA extracted from cytosol fractions following exonuclease treatment results in highest mtDNA yield and purity from frozen tumor tissue (270-fold mtDNA enrichment). Next, we demonstrate the sensitivity of detection of low-frequent single-nucleotide mtDNA variants (≤1% allele frequency) in breast cancer cell lines MDA-MB-231 and MCF-7 by single-molecule real-time (SMRT) sequencing, UltraSEEK chemistry based mass spectrometry, and digital PCR. We also show de novo detection and allelic phasing of variants by SMRT sequencing. We conclude that our sensitive procedure to detect low-frequent single-nucleotide mtDNA variants from frozen tumor tissue is based on extraction of DNA from cytosol fractions followed by exonuclease treatment to obtain high mtDNA purity, and subsequent SMRT sequencing for (de novo) detection and allelic phasing of variants.

AAV-mediated in vivo knockdown of luciferase using combinatorial RNAi and U1i.

RNA interference (RNAi) has been successfully employed for specific inhibition of gene expression; however, safety and delivery of RNAi remain critical issues. We investigated the combinatorial use of RNAi and U1 interference (U1i). U1i is a gene-silencing technique that acts on the pre-mRNA by preventing polyadenylation. RNAi and U1i have distinct mechanisms of action in different cellular compartments and their combined effect allows usage of minimal doses, thereby avoiding toxicity while retaining high target inhibition. As a proof of concept, we investigated knockdown of the firefly luciferase reporter gene by combinatorial use of RNAi and U1i, and evaluated their inhibitory potential both in vitro and in vivo. Co-transfection of RNAi and U1i constructs showed additive reduction of luciferase expression up to 95% in vitro. We attained similar knockdown when RNAi and U1i constructs were hydrodynamically transfected into murine liver, demonstrating for the first time successful in vivo application of U1i. Moreover, we demonstrated long-term gene silencing by AAV-mediated transduction of murine muscle with RNAi/U1i constructs targeting firefly luciferase. In conclusion, these results provide a proof of principle for the combinatorial use of RNAi and U1i to enhance target gene knockdown in vivo.

Strong-coupling polarons in dilute gas Bose-Einstein condensates.

A neutral impurity atom immersed in a dilute Bose-Einstein condensate (BEC) can have a bound ground state in which the impurity is self-localized. In this polaronlike state, the impurity distorts the density of the surrounding BEC, thereby creating the self-trapping potential minimum. We describe the self-localization in a strong-coupling approach.

Degenerate fermion gas heating by hole creation.

Loss processes that remove particles from an atom trap leave holes behind in the single particle distribution if the trapped gas is a degenerate fermion system. The appearance of holes increases the temperature and we show that the heating (i) is significant if the initial temperature is well below the Fermi temperature T(F), and (ii) increases the temperature to T> or =T(F)/4 after half of the system's lifetime, regardless of the initial temperature. The hole heating has important consequences for the prospect of observing Cooper pairing in atom traps.

One-tube real-time isothermal amplification assay to identify and distinguish human immunodeficiency virus type 1 subtypes A, B, and C and circulating recombinant forms AE and AG.

To halt the human immunodeficiency virus type 1 (HIV-1) epidemic requires interventions that can prevent transmission of numerous HIV-1 subtypes. The most frequently transmitted viruses belong to the subtypes A, B, and C and the circulating recombinant forms (CRFs) AE and AG. A fast one-tube assay that identifies and distinguishes among subtypes A, B, and C and CRFs AE and AG of HIV-1 was developed. The assay amplifies a part of the gag gene sequence of the genome of all currently known HIV-1 subtypes and can identify and distinguish among the targeted subtypes as the reaction proceeds, because of the addition of subtype-specific molecular beacons with multiple fluorophores. The combination of isothermal nucleic acid sequence-based amplification and molecular beacons is a new approach in the design of real-time assays. To obtain a sufficiently specific assay, we developed a new strategy in the design of molecular beacons, purposely introducing mismatches in the molecular beacons. The subtype A and CRF AG isolates reacted with the same molecular beacon. We tested the specificity and sensitivity of the assay on a panel of the culture supernatant of 34 viruses encompassing all HIV-1 subtypes: subtypes A through G, CRF AE and AG, a group O isolate, and a group N isolate. Assay sensitivity on this panel was 92%, with 89% correct subtype identification relative to sequence analysis. A linear relationship was found between the amount of input RNA in the reaction mixture and the time that the reaction became positive. The lower detection level of the assay was approximately 10(3) copies of HIV-1 RNA per reaction. In 38% of 50 serum samples from HIV-1-infected individuals with a detectable amount of virus, we could identify subtype sequences with a specificity of 94% by using sequencing and phylogenetic analysis as the "gold standard." In conclusion, we showed the feasibility of the approach of using multiple molecular beacons labeled with different fluorophores in combination with isothermal amplification to identify and distinguish subtypes A, B, and C and CRFs AE and AG of HIV-1. Because of the low sensitivity, the assay in this format would not be suited for clinical use but can possibly be used for epidemiological monitoring as well as vaccine research studies.

Driving superfluidity with photoassociation.

We theoretically examine photoassociation of a two-component Fermi degenerate gas. Our focus is on adjusting the atom-atom interaction, and thereby increasing the critical temperature of the BCS transition to the superfluid state. In order to avoid spontaneous decay of the molecules, the photoassociating light must be far-off resonance. Very high light intensities are therefore required for effective control of the BCS transition.

Scalability, reliability, and validity of the benzodiazepine dependence self-report questionnaire in outpatient benzodiazepine users.

As there is no multidimensional instrument available that reflects the severity of benzodiazepine (BZD) dependence comprehensively, the Benzodiazepine Dependence Self-Report Questionnaire (Bendep-SRQ) was developed and investigated. The Bendep-SRQ, Symptom Checklist-90 (SCL-90), Schedules for Clinical Assessments in Neuropsychiatry (SCAN), and Addiction Severity Index-Revised (ASI-R) were administered to 115 general practice (GP) patients, 124 psychiatric outpatients, and 33 self-help patients who were using BZDs. Factor and Rasch analyses were applied to construct scales. Reliability assessments were made in terms of subject discriminability, item discriminability, and test-retest stability. To support the construct validity of the scales, theoretical rationales were required to explain the specific item order provided by the Rasch scale values. To assess the concurrent and discriminant validity, a matrix consisting of the above-mentioned measures was factor-analyzed. Four Rasch-homogeneous scales were delineated: problematic use, preoccupation, lack of compliance, and withdrawal. Nearly all subject discriminability, item discriminability, and test-retest results indicated good reliability. A BZD dependence factor was extracted with high loadings for the Bendep-SRQ scales and the concurrent measures. The discriminant measures had high loadings on other factors. The scalability, reliability, and validity of the Bendep-SRQ scales appeared to be good. The Bendep-SRQ shows great promise as a useful and easily manageable instrument for assessment of the severity of BZD dependence in clinical practice and scientific research.